TOBACCO INBRED PLANTS K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC AND HYBRID NC196 SRC

INCORPORATION OF SEQUENCE LISTING

The Sequence Listing is hereby incorporated by reference in its entirety, including the file named P34035US00.txt, which is 38,002 bytes in size and was created on Jan. 10, 2013, which is likewise herein incorporated by reference in its entirety.

FIELD

The present disclosure provides tobacco inbred plants K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC and hybrid cultivar NC196 SRC. The present disclosure also provides parts of such plants and products made from those parts. The present disclosure also includes progeny of the provided plants including hybrids.

BACKGROUND

Tobacco (Nicotiana tabacum L.) is an important commercial crop in the United States as well as in other countries. In tobacco plants, N-demethylation of nicotine results in nornicotine, a secondary alkaloid known to be a precursor for formation of N-Nitrosonornicotine (“NNN”) in cured leaves. NNN is an undesired component of cured leaves.

The predominant alkaloid found in commercial tobacco varieties is nicotine, typically accounting for 90-95% of the total alkaloid pool. The remaining alkaloid fraction is comprised primarily of three additional pyridine alkaloids: nornicotine, anabasine, and anatabine. Nornicotine is generated directly from nicotine through the activity of the enzyme nicotine N-demethylase. Nornicotine usually represents less than 5% of the total pyridine alkaloid pool, but through a process termed “conversion,” tobacco plants that initially produce very low amounts of nornicotine give rise to progeny that metabolically “convert” a large percentage of leaf nicotine to nornicotine. In tobacco plants that have genetically converted (termed “converters”), the great majority of nornicotine production occurs during the senescence and curing of the mature leaf (Wernsman and Matzinger (1968), Tob. Sci., 12:226-228). Burley tobaccos are particularly prone to genetic conversion, with rates as high as 20% per generation observed in some cultivars.

During the curing and processing of the tobacco leaf, a portion of the nornicotine is metabolized to the compound NNN, a tobacco-specific nitrosamine (TSNA) that has been asserted to be carcinogenic in laboratory animals (Hecht and Hoffmann (1990), Cancer Surveys, 8:273-294; Hoffmann et al, (1994), J. Toxicol. Environ. Health, 41:1-52; Hecht (1998), Chem. Res. Toxicol., 11:559-603). In flue-cured tobaccos, TSNAs are found to be predominantly formed through the reaction of alkaloids with the minute amounts of nitrogen oxides present in combustion gases formed by the direct-fired heating systems found in traditional curing barns (Peele and Gentry (1999), “Formation of Tobacco-specific Nitrosamines in Flue-cured Tobacco,” CORESTA Meeting, Agro-Phyto Groups, Suzhou, China). Retrofitting these curing barns with heat-exchangers virtually eliminated the mixing of combustion gases with the curing air and dramatically reduced the formation of TSNAs in tobaccos cured in this manner (Boyette and Hamm (2001), Rec. Adv. Tob. Sci., 27:17-22.). In contrast, in the air-cured Burley tobaccos, TSNA formation proceeds primarily through reaction of tobacco alkaloids with nitrite, a process catalyzed by leaf-borne microbes (Bush et al. (2001), Rec. Adv. Tob. Sci., 27:23-46). Thus far, attempts to reduce TSNAs through modification of curing conditions while maintaining acceptable quality standards have not proven to be successful for the air-cured tobaccos.

SUMMARY

In an aspect, the present disclosure includes a seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455.

In another aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455.

In a further aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455.

In an aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a harvested leaf, or part thereof, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN), where the reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN) is reduced in a smoke stream produced from the leaf.

In a further aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a part of a tobacco plant, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the part is selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod and petiole.

In another aspect, the present disclosure includes a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In an aspect, the present disclosure includes a tobacco plant regenerated from a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole where the regenerated plant has all, or essentially all of the morphological and physiological characteristics of cultivar K326 SRC.

In an aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455.

In another aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where the F₁plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes an F₁progeny seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455.

In an aspect, the present disclosure also includes a container of F₁progeny seeds produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure includes an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, and where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure further includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN) produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) in a smoke stream produced from the leaf, where the plant is produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In an aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In another aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In an aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In another aspect, the present disclosure includes a harvested leaf, or part thereof, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN), where the reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN) is reduced in a smoke stream produced from the leaf.

In a further aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a part of a tobacco plant, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the part is selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In another aspect, the present disclosure includes a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In an aspect, the present disclosure includes a tobacco plant regenerated from a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole where the regenerated plant has all, or essentially all of the morphological and physiological characteristics of cultivar CMS K326 SRC.

In an aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In another aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, where the F₁plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes an F₁progeny seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a plant of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13454, where the plant of tobacco cultivar CMS K326 SRC is the female parent.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a plant of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In an aspect, the present disclosure also includes a container of F₁progeny seeds produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In an aspect, the present disclosure includes an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In another aspect, the present disclosure further includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN) produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) in a smoke stream produced from the leaf, where the plant is produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In an aspect, the present disclosure includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, and where the tobacco product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and further where the product has a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN).

In an aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile, where the cytoplasmic male sterile plant is a plant of tobacco cultivar CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454.

In another aspect, the present disclosure includes a seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453.

In an aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453.

In another aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453.

In an aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In another aspect, the present disclosure includes a harvested leaf, or part thereof, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN), where the reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) is reduced in a smoke stream produced from the leaf.

In a further aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a part of a tobacco plant, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the part is selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In another aspect, the present disclosure includes a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In an aspect, the present disclosure includes a tobacco plant regenerated from a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole where the regenerated plant has all, or essentially all of the morphological and physiological characteristics of cultivar K346 SRC.

In an aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453.

In another aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the F₁plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the plant of tobacco cultivar K346 SRC is the male parent.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where the plant of tobacco cultivar K346 SRC is the female parent.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes an F₁progeny seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a container of F₁progeny seeds produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure includes an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, and where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure further includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) in a smoke stream produced from the leaf, where the plant is produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and further where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In another aspect, the present disclosure includes a seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In an aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In another aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In an aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In another aspect, the present disclosure includes a harvested leaf, or part thereof, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN), where the reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) is reduced in a smoke stream produced from the leaf.

In a further aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a part of a tobacco plant, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the part is selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In another aspect, the present disclosure includes a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In an aspect, the present disclosure includes a tobacco plant regenerated from a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole where the regenerated plant has all, or essentially all of the morphological and physiological characteristics of cultivar CMS K346 SRC.

In an aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession PTA-13452.

In another aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, where the F₁plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes an F₁progeny seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a plant of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13452, where the plant of tobacco cultivar CMS K346 SRC is the female parent.

In an aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile, where the cytoplasmic male sterile plant is a plant of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In an aspect, the present disclosure also includes a container of F₁progeny seeds produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In an aspect, the present disclosure includes an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In another aspect, the present disclosure further includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) in a smoke stream produced from the leaf, where the plant is produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452.

In an aspect, the present disclosure includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, and where the tobacco product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and further where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457.

In another aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457.

In a further aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457.

In an aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN).

In an aspect, the present disclosure includes a harvested leaf, or part thereof, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN), where the reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) is reduced in a smoke stream produced from the leaf.

In a further aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a part of a tobacco plant, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the part is selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In another aspect, the present disclosure includes a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In an aspect, the present disclosure includes a tobacco plant regenerated from a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole where the regenerated plant has all, or essentially all of the morphological and physiological characteristics of cultivar NC1562-1 SRC.

In an aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457.

In another aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the F₁plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes an F₁progeny seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the plant of tobacco cultivar NC1562-1 SRC is the female parent.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where the plant of tobacco cultivar NC1562-1 SRC is the male parent.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a container of F₁progeny seeds produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure includes an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, and where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure further includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN) in a smoke stream produced from the leaf, where the plant is produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and further where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314.

In another aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314.

In a further aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314.

In an aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a harvested leaf, or part thereof, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN), where the reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) is reduced in a smoke stream produced from the leaf.

In a further aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a part of a tobacco plant, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the part is selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In another aspect, the present disclosure includes a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In an aspect, the present disclosure includes a tobacco plant regenerated from a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole where the regenerated plant has all, or essentially all of the morphological and physiological characteristics of cultivar NCTG-61 SRC.

In an aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314.

In another aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the F₁plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes an F₁progeny seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the plant of tobacco cultivar NCTG-61 SRC is the female parent.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where the plant of tobacco cultivar NCTG-61 SRC is the male parent.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-120314, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a container of F₁progeny seeds produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure includes an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure further includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN) produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN) in a smoke stream produced from the leaf, where the plant is produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where at least one tobacco plant is cytoplasmic male sterile (CMS).

In an aspect, the present disclosure includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is produced by growing a seed of tobacco cultivar NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-120314, where at least one tobacco plant is cytoplasmic male sterile, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and further where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In another aspect, the present disclosure includes a seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In an aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In another aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In an aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In another aspect, the present disclosure includes a harvested leaf, or part thereof, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN), where the reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) is reduced in a smoke stream produced from the leaf.

In a further aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a part of a tobacco plant, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the part is selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In another aspect, the present disclosure includes a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In an aspect, the present disclosure includes a tobacco plant regenerated from a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole where the regenerated plant has all, or essentially all of the morphological and physiological characteristics of cultivar CMS NCTG-61 SRC.

In an aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In another aspect, the present disclosure includes an F₁progeny plant of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the F₁plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes an F₁progeny seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a plant of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In another aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, where the plant of tobacco cultivar CMS NCTG-61 SRC is the female parent.

In an aspect, the present disclosure includes a method for producing a tobacco seed comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile, where the cytoplasmic male sterile plant is a plant of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In an aspect, the present disclosure also includes a container of F₁progeny seeds produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In an aspect, the present disclosure includes an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In another aspect, the present disclosure further includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonomicotine (NNN) produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In an aspect, the present disclosure also includes a harvested leaf of an F₁progeny plant having a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) in a smoke stream produced from the leaf, where the plant is produced by growing a seed produced by a method of comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456.

In an aspect, the present disclosure includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and where the tobacco product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In an aspect, the present disclosure further includes a tobacco product prepared from an F₁progeny tobacco plant, or part thereof, where the plant or part thereof is produced by growing a F₁progeny seed produced by a method comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where at least one tobacco plant is cytoplasmic male sterile (CMS), where the cytoplasmic male sterile plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and where the tobacco product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco, and further where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In another aspect, the present disclosure includes a seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458.

In an aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing a seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458.

In a further aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, or part thereof, produced by growing the seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458.

In another aspect, the present disclosure includes a harvested leaf, or part thereof, of a tobacco plant, produced by growing the seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a harvested leaf, or part thereof, produced by growing the seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, where the leaf has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN), where the reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN) is reduced in a smoke stream produced from the leaf.

In a further aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, where the product is selected from the group consisting of pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco.

In another aspect, the present disclosure includes a tobacco product, prepared from a tobacco plant, or part thereof, produced by growing the seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, where the product is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, and chewing tobacco where the product has a reduced amount of nornicotine and/or N′-nitrosonornicotine (NNN).

In an aspect, the present disclosure includes a part of a tobacco plant, produced by growing a seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, where the part is selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In a further aspect, the present disclosure includes a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13458, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole.

In an aspect, the present disclosure includes a tobacco plant regenerated from a tissue culture produced from a protoplast or cell from a tobacco plant, or part thereof, produced by growing the seed of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, where the cell or protoplast of the tissue culture is produced from a plant part selected from the group consisting of a leaf, pollen, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, shoot, stem, pod, and petiole, where the regenerated plant has all, or essentially all of the morphological and physiological characteristics of hybrid cultivar NC196 SRC.

In an aspect, the present disclosure includes an F₁progeny plant of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458.

In another aspect, the present disclosure includes an F₁progeny plant of hybrid tobacco cultivar NC196 SRC, a representative sample seed of the hybrid cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, where the F₁plant is cytoplasmic male sterile (CMS).

In another aspect, the present disclosure includes a method for producing a tobacco seed of NC196 SRC comprising crossing two tobacco plants and harvesting the resultant tobacco seed, where one tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13456 and a second tobacco plant is a tobacco plant produced by growing the seed of tobacco cultivar NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457.

In an aspect, the present disclosure includes a method of vegetatively propagating a plant of a tobacco cultivar comprising the steps of (a) collecting tissue capable of being propagated from a plant of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458; (b) cultivating the tissue to obtain a proliferated shoot; and (c) rooting the proliferated shoots to obtain a rooted plantlet.

In an aspect, the present disclosure includes a method of vegetatively propagating a plant of a tobacco cultivar comprising the steps of (a) collecting tissue capable of being propagated from a plant of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458; (b) cultivating the tissue to obtain a proliferated shoot; (c) rooting the proliferated shoots to obtain a rooted plantlet; and (d) growing a plant from the rooted plantlet.

In an aspect, the present disclosure includes a method of introducing a desired trait into a tobacco cultivar comprising: (a) crossing a plant of a first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, with a second tobacco plant that comprises a desired trait to produce an F₁progeny seed; (b) growing the F₁progeny seed and selecting an F₁progeny plant that comprises the desired trait; (c) crossing the selected F₁progeny plant with a plant of said first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456 to produce a backcross BC₁F₁progeny seed; (d) growing the BC₁F₁progeny seed and selecting a backcross BC₁F₁progeny plant comprising the desired trait and essentially all of the physiological and morphological characteristics of said first tobacco cultivar, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC; and (e) repeating steps (c) and (d) three or more times in succession to produce selected fourth or higher backcross progeny that comprise the desired trait.

In an aspect, the present disclosure includes a method of introducing a desired trait into a tobacco cultivar comprising: (a) crossing a plant of a first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, with a second tobacco plant that comprises a desired trait to produce an F₁progeny seed; (b) growing the F₁progeny seed and selecting an F₁progeny plant that comprises the desired trait; (c) crossing the selected F₁progeny plant with a plant of the first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, and NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456 to produce a backcross BC₁F₁progeny seed; (d) growing the BC₁F₁F₂progeny seed and selecting a backcross BC₁F₁progeny plant comprising the desired trait and essentially all of the physiological and morphological characteristics of the first tobacco cultivar, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC; and (e) repeating steps (c) and (d) three or more times in succession to produce selected fourth or higher backcross progeny that comprise the desired trait, where the trait is cytoplasmic male sterility (CMS).

In an aspect, the present disclosure includes a method of introducing a desired trait into a tobacco cultivar comprising: (a) crossing a plant of a first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, with a second tobacco plant that comprises a desired trait to produce an F₁progeny seed; (b) growing the F₁progeny seed and selecting an F₁progeny plant that comprises the desired trait; (c) crossing the selected F₁progeny plant with a plant of the first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457 to produce a backcross BC₁F₁progeny seed; (d) growing the BC₁F₁progeny seed and selecting a backcross BC₁F₁progeny plant comprising the desired trait and the physiological and essentially all of morphological characteristics of the first tobacco cultivar K326 SRC, K346 SRC, NCTG-61 SRC, or NC1562-1 SRC; and (e) repeating steps (c) and (d) three or more times in succession to produce selected fourth or higher backcross progeny that comprise the desired trait, where the trait is cytoplasmic male sterility (CMS) and the CMS trait is obtained from the cytoplasm of Nicotiana suaveolens or Nicotiana glauca.

In an aspect, the present disclosure includes a tobacco plant produced by a method comprising introducing a desired trait into a tobacco cultivar comprising: (a) crossing a plant of a first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, with a second tobacco plant that comprises a desired trait to produce an F₁progeny seed; (b) growing the F₁progeny seed and selecting an F₁progeny plant that comprises the desired trait; (c) crossing the selected F₁progeny plant with a plant of the first tobacco cultivar selected from the group consisting of K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, and CMS NCTG-61 SRC to produce a backcross BC₁F₁progeny seed; (d) growing the BC₁F₁progeny seed and selecting a backcross BC₁F₁progeny plant comprising the desired trait and essentially all of the physiological and morphological characteristics of the first tobacco cultivar K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC; and (e) repeating steps (c) and (d) three or more times in succession to produce selected fourth or higher backcross progeny that comprise the desired trait.

In another aspect, the present disclosure includes a method of introducing a desired trait into a tobacco cultivar comprising: (a) crossing a plant of a first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, with a plant of a second tobacco cultivar that comprises a desired trait to produce a progeny plant where the desired trait is selected from the group consisting of herbicide resistance, pest resistance, disease resistance, high yield, high grade index, curability, curing quality, mechanical harvestability, holding ability, leaf quality, height, plant maturation, early maturing, early to medium maturing, medium maturing, medium to late maturing, late maturing; small stalk, medium stalk, large stalk, leaf number per plant, 5-10 leaves per plant, 11-15 leaves per plant, 16-21 leaves per plant, and any combination thereof, to produce an F₁progeny seed; (b) growing the F₁progeny seed into an F₁progeny plant and selecting the F₁progeny plant having the desired trait; (c) crossing the selected F₁progeny plant with a plant of the first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456 to produce a backcross progeny plant seed; (d) growing the backcross progeny plant seed into a backcross progeny plant and selecting the backcross progeny plant comprising the desired trait and essentially all of the physiological and morphological characteristics of the first tobacco cultivar, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC; and (e) repeating steps (c) and (d) one or more times in succession to produce a selected fourth or higher backcross progeny plant that comprises the desired trait and essentially all of the physiological and morphological characteristics of the first tobacco cultivar, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC.

In another aspect, the present disclosure includes a method of introducing a desired trait into a tobacco cultivar comprising: (a) crossing a plant of a first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, with a plant of a second tobacco cultivar that comprises a desired trait to produce a progeny plant where the desired trait is selected from the group consisting of herbicide resistance, pest resistance, disease resistance, high yield, high grade index, curability, curing quality, mechanical harvestability, holding ability, leaf quality, height, plant maturation, early maturing, early to medium maturing, medium maturing, medium to late maturing, late maturing; small stalk, medium stalk, large stalk, leaf number per plant, 5-10 leaves per plant, 11-15 leaves per plant, 16-21 leaves per plant, and any combination thereof, to produce an F₁progeny seed; (b) growing the F₁progeny seed into an F₁progeny plant and selecting the F₁progeny plant having the desired trait; (c) crossing the selected F₁progeny plant with a plant of the first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456 to produce a backcross progeny plant seed; (d) growing the backcross progeny plant seed into a backcross progeny plant and selecting the backcross progeny plant comprising the desired trait and essentially all of the physiological and morphological characteristics of a tobacco cultivar, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC; and (e) repeating steps (c) and (d) one or more times in succession to produce a selected fourth or higher backcross progeny plant that comprises the desired trait and essentially all of the physiological and morphological characteristics of the first tobacco cultivar, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC, where the plant has a desired trait.

In another aspect, the present disclosure includes a method of introducing a desired trait into a tobacco cultivar comprising: (a) crossing a plant of a first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, with a plant of a second tobacco cultivar that comprises a desired trait to produce a progeny plant where the desired trait is selected from the group consisting of herbicide resistance, pest resistance, disease resistance, high yield, high grade index, curability, curing quality, mechanical harvestability, holding ability, leaf quality, height, plant maturation, early maturing, early to medium maturing, medium maturing, medium to late maturing, late maturing; small stalk, medium stalk, large stalk, leaf number per plant, 5-10 leaves per plant, 11-15 leaves per plant, 16-21 leaves per plant, and any combination thereof, to produce an F₁progeny seed; (b) growing the F₁progeny seed into an F₁progeny plant and selecting the F₁progeny plant having the desired trait; (c) crossing the selected F₁progeny plant with a plant of the first tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456 to produce a backcross progeny plant seed; (d) growing the backcross progeny plant seed into a backcross progeny plant and selecting the backcross progeny plant comprising the desired trait and essentially all of the physiological and morphological characteristics of the first tobacco cultivar, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC; and (e) repeating steps (c) and (d) one or more times in succession to produce a selected fourth or higher backcross progeny plant that comprises the desired trait and essentially all of the physiological and morphological characteristics of a tobacco cultivar, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, or CMS NCTG-61 SRC, where the plant has a desired trait of disease resistance.

In another aspect, the present disclosure includes a method for producing a tobacco plant having decreased nicotine conversion comprising: identifying a first tobacco plant having the sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 11, and combinations thereof; crossing the first tobacco plant with a second tobacco plant and collecting an F₁seed; crossing a plant grown from the F₁seed to a third tobacco plant and collecting a second tobacco seed; and identifying a second tobacco seed or a plant grown from the second seed that is homozygous for the sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 11, and combinations thereof.

In an aspect, the present disclosure includes a method for producing a tobacco plant having decreased nicotine conversion comprising: identifying a first tobacco plant having a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 11, and combinations thereof; crossing the first tobacco plant with a second tobacco plant and collecting an F₁seed; crossing a plant grown from the F₁seed to a third tobacco plant and collecting a second tobacco seed; and identifying a second tobacco seed or a plant grown from the second seed that is homozygous for a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 11, and combinations thereof, where the first tobacco plant is a plant of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458.

In an aspect, the present disclosure includes a method for producing a tobacco plant having decreased nicotine conversion comprising: identifying a first tobacco plant having a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 11, and combinations thereof; crossing the first tobacco plant with a second tobacco plant and collecting an F₁seed; crossing a plant grown from the F₁seed to a third tobacco plant and collecting a second tobacco seed; and identifying a second tobacco seed or a plant grown from the second seed that is homozygous for a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 11, and combinations thereof, where the third tobacco plant is a plant of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13458.

In another aspect, the present disclosure includes a method of producing a plant of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456 having an additional desired trait comprising the steps of: (a) collecting tissue capable of being propagated from a plant of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-120314, and CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456; and (b) introducing a transgene conferring the desired trait into the tissue.

In another aspect, the present disclosure includes a method of producing an herbicide resistant tobacco plant comprising transforming a tobacco plant, or part thereof, produced by growing a seed of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458 with a transgene where the transgene confers resistance to an herbicide selected from the group consisting of imidazolinone, cyclohexanedione, sulfonylurea, glyphosate, glufosinate, phenoxy proprionic acid, L-phosphinothricin, triazine, and benzonitrile.

In another aspect, the present disclosure includes an herbicide resistant tobacco plant produced by a method comprising transforming a tobacco plant, or part thereof, produced by growing a seed of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458 with a transgene where the transgene confers resistance to an herbicide selected from the group consisting of imidazolinone, cyclohexanedione, sulfonylurea, glyphosate, glufosinate, phenoxy proprionic acid, L-phosphinothricin, triazine, and benzonitrile.

In another aspect, the present disclosure includes a method of producing a pest or insect resistant tobacco plant where the method comprises transforming a tobacco plant produced by growing a seed of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458 with a transgene that confers pest or insect resistance.

In a further aspect, the present disclosure includes a pest or insect resistant tobacco plant produced by a method comprising transforming a tobacco plant produced by growing a seed of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458 with a transgene that confers pest or insect resistance.

In a further aspect, the present disclosure includes a pest or insect resistant tobacco plant produced by a method comprising transforming a tobacco plant produced by growing a seed of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458, with a transgene that confers pest or insect resistance, where the transgene encodes a Bacillus thuringiensis (BT) endotoxin.

In another aspect, the present disclosure includes a method of producing a disease resistant tobacco plant where the method comprises transforming a tobacco plant produced by growing a seed of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458 with a transgene that confers disease resistance.

In a further aspect, the present disclosure includes a disease resistant tobacco plant produced by a method comprising transforming a tobacco plant produced by growing a seed of a tobacco cultivar selected from the group consisting of K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455, CMS K326 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454, K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453, CMS K346 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452, NC1562-1 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13457, NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314, CMS NCTG-61 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456, and hybrid cultivar NC196 SRC, a representative sample seed of the cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458 with a transgene that confers disease resistance.

BRIEF DESCRIPTION OF THE SEQUENCES

SEQ ID NO: 1 sets forth a cyp82e4 W329Stop nucleotide sequence.

SEQ ID NO: 2 sets forth a cyp82e5v2 W422Stop nucleotide sequence.

SEQ ID NO: 3 sets forth a cyp82e4 W329Stop amino acid sequence.

SEQ ID NO: 4 sets forth a cyp82e5v2 W422Stop amino acid sequence.

SEQ ID NO: 5 sets forth a CYP82E4 wild-type nucleotide sequence.

SEQ ID NO: 6 sets forth a CYP82E5v2 wild-type nucleotide sequence.

SEQ ID NO: 7 sets forth a CYP82E4 wild-type amino acid sequence.

SEQ ID NO: 8 sets forth a CYP82E5v2 wild-type amino acid sequence.

SEQ ID NO: 9 sets forth a CYP82E10 wild-type nucleotide sequence.

SEQ ID NO: 10 sets forth a CYP82E10 wild-type amino acid sequence.

SEQ ID NO: 11 sets forth a CYP82E10 P381S nucleotide sequence.

SEQ ID NO: 12 sets forth a CYP82E10 P381S amino acid sequence.

All three tobacco Nicotine Demethylase genes (CYP82E4, CYP82E5v2, CYP82E10) share a common structure: a 939 bp exon 1 and a 612 bp exon 2 separated by a large intron, whose length varies among the three genes. See Lewis et al., “Three nicotine demethylase genes mediate nornicotine biosynthesis in Nicotiana tabacum L.: Functional characterization of the CYP82E10 gene,” Phytochemistry, 71 (2010), 1988-1998. SEQ ID NOs: 1, 2, 5, 6, 9, and 11 set forth wild-type or mutant versions of coding sequences of CYP82E4, CYP82E5v2, and CYP82E10. It is understood that, used herein, a plant comprising, having, or homozygous for a sequence selected from SEQ ID NOs: 1, 2, 5, 6, 9, and 11 refers to a plant comprising at the CYP82E4, CYP82E5v2, or CYP82E10 endogenous locus a genomic sequence comprising the coding sequence of SEQ ID NO: 1, 2, 5, 6, 9, or 11.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1: Comparisons between K326 SRC (shown as “K 326 e4e5e10 Mutant”) and K326 for their NNN concentrations in tobacco filler and cigarette smoke, and their percent nicotine conversions in tobacco filler. K326 SRC has a lower level of NNN compared to K326 in both tobacco filler (˜0.22 versus ˜0.53 ppm, shown as “Filler NNN”) and cigarette smoke (˜0.015 versus ˜0.04 ng/mg Total Particle Matter (TPM), shown as “Smoke NNN”). Percent nicotine conversion is lower in K326 SRC tobacco filler (˜0.8%) than K326 tobacco filler (˜3.1%, shown as “Filler % Conversion”).

DETAILED DESCRIPTION

Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. One skilled in the art will recognize many methods can be used in the practice of the present disclosure. Indeed, the present disclosure is in no way limited to the methods and materials described. For purposes of the present disclosure, the following terms are defined below.

Any references cited herein are incorporated by reference in their entireties.

As used herein, the singular form “a,” “an,’ and “the” include plural references unless the context clearly dictates otherwise. For example, the term “a compound” or “at least one compound” may include a plurality of compounds, including mixtures thereof.

The term “about” is used herein to mean approximately, roughly, around, or in the region of. When the term “about” is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the numerical values set forth.

“Tobacco product” is defined as “any product made or derived from tobacco that is intended for human use or consumption, including any component, part, or accessory of a tobacco product (except for raw materials other than tobacco used in manufacturing a component, part, or accessory of a tobacco product)” (section 201 of the FD&C Act; 21 U.S.C. 321). The label and packaging is part of a tobacco product.

Terms “nicotine conversion rate,” “percent nicotine conversion,” and “percentage nicotine conversion” are used interchangeably. Percent nicotine demethylation in a sample is calculated by dividing the level of nornicotine by the combined level of nicotine and nornicotine as measured in the sample, and multiplying by 100.

K326 SRC

The present disclosure also provides tobacco cultivars, and parts thereof, from K326 SRC, representative sample seeds of this cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455. The present disclosure also includes a tobacco plant, or part thereof, produced by growing a seed of K326 SRC. A plant of the present disclosure can further include a plant with all, or essentially all of the morphological and physiological characteristics of cultivar K326 SRC.

While not being limited by process, K326 SRC is a result of the introduction of three mutated CYP82 genes in a flue-cured tobacco cultivar K326. The three genes are a mutated CYP82E4 gene recited as 325-6 #775 in Lewis et al. (“Three nicotine demethylase genes mediate nornicotine biosynthesis in Nicotiana tabacum L.: Functional characterization of the CYP82E10 gene,” Phytochemistry, 71 (2010), 1988-1998 (SEQ ID NO: 1, which sets forth a cyp82e4 W329Stop, hereby incorporated by reference in its entirety)), a mutated CYP82E5v2 recited in Lewis et al. (supra) as 325-6 #1-13 (SEQ ID NO: 2, which sets forth a cyp82e5v2 W422Stop, hereby incorporated by reference in its entirety), and a mutated CYP82E10 recited in Lewis et al. (supra) as 325-6 #1041 (SEQ ID NO:11, which sets forth a cyp82e10 P381S, hereby incorporated by reference in its entirety). Mutations cyp82e4 W329Stop and cyp82e5v2 W422Stop result in truncated proteins while cyp82E10 P381S results in a nonfunctional protein. A cyp82e4 W329Stop (“e4”), a cyp82e5v2 W422Stop (“e5”), and a cyp82e10 P381S (“e10”) mutation are introduced from a e4e5|e4e5|e10e10 triple mutant from a strong converter burley background, line DH98-325-6, as listed in Table 2 of Lewis et al. (supra) into a flue-cured tobacco cultivar K326 background.

K326 SRC is the result of seven backcrosses with flue-cured cultivar K326 as the recurrent parent, followed by two rounds of selling with selection for homozygosity for the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S mutations to yield BC₇F₃plants (K326 SRC) in which the wild-type CYP82E4, CYP82E5v2 and CYP82E10 alleles of K326 are replaced by the mutant (e.g., cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S) alleles.

K326 SRC progeny plants have genetic backgrounds that are at least 95%, at least 97%, at least 98%, or at least 99% similar to K326. K326 SRC plants exhibit low nornicotine levels and produce leaves with reduced potential for accumulating derived NNN during curing, storage, and smoking.

CMS K326 SRC

The present disclosure also provides tobacco cultivars, and parts thereof, from CMS K326 SRC, representative sample seeds of this cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454. The present disclosure also includes a tobacco plant, or part thereof, produced by growing a seed of CMS K326 SRC. A plant of the present disclosure can further include a plant with all, or essentially all, of the morphological and physiological characteristics of cultivar CMS K326 SRC. CMS K326 SRC is a male-sterile (CMS) version of K326 SRC (CMS K326SRC) produced by crossing a plant of CMS K326 as a female with pollen of K326 SRC BC₆F₁to produce male-sterile plants heterozygous for all three mutations. Progeny plants of the CMS K326×K326 SRC BC₆F₁cross are male sterile. A plurality of CMS K326×K326 SRC BC₆F₁×CMS plants (e.g., CMS F₁progeny plants) are screened for the cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S mutations to identify plants heterozygous for all three mutations. A single male-sterile plant resulting from this cross is backcrossed as a female to K326 SRC to prepare BC₇F₁CMS progeny. BC₇F₁CMS progeny homozygous for the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S mutations are identified by genotyping and designated as CMS K326 SRC. Because the CMS K326 SRC line is male sterile, it is maintained via pollination with K326 SRC. K326 SRC is crossed as the male parent to CMS K326 SRC to prepare CMS K326 SRC F₁progeny plants.

CMS K326 SRC and CMS K326 SRC F₁progeny plants have genetic backgrounds that are at least 95%, at least 97%, at least 98%, or at least 99% similar to K326. CMS K326 SRC and CMS K326 SRC F₁progeny plants exhibit low nornicotine levels and produce leaves with reduced potential for accumulating derived NNN during curing, storage, and smoking.

K346 SRC

The present disclosure also provides tobacco cultivars, and parts thereof, from K346 SRC, where representative sample seeds of this cultivar have been deposited with the ATCC under ATCC Accession No. PTA-13453. The present disclosure also includes a tobacco plant, or part thereof, produced by growing a seed of K346 SRC. A plant of the present disclosure can include a plant with all, or essentially all, of the morphological and physiological characteristics of cultivar K346 SRC. While not being limited by process, K346 SRC is a result of introducing the cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S mutations from DH98-325-6. F₁individuals originating from a cross between K346 and DH98-325-6 and heterozygous for each mutation are then backcrossed seven times to K346 to produce BC₇F₁progeny. BC₇F₁individuals heterozygous for all three mutations are self-pollinated to produce BC₇F₂seed and individuals homozygous for all three mutations identified. A single BC₇F₂plant is self-pollinated to produce BC₇F₃(K346 SRC) in which the wild-type CYP82E4, CYP82E5v2 and CYP82E10 alleles of K346 are replaced by the mutant (e.g., cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S).

K346 SRC has a genetic background that is at least 95%, at least 97%, at least 98%, or at least 99% similar to K346. K346 SRC exhibits low nornicotine levels and produce leaves with reduced potential for accumulating derived NNN during curing, storage, and smoking.

CMS K346 SRC

The present disclosure also provides tobacco cultivars, and parts thereof, from CMS K346 SRC, representative sample seeds of this cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452. The present disclosure also includes a tobacco plant, or part thereof, produced by growing a seed of CMS K346 SRC. A plant of the present disclosure can further include a plant with all, or essentially all, of the morphological and physiological characteristics of cultivar CMS K426 SRC. CMS K346 SRC is a male-sterile (CMS) version of K346 SRC (CMS K346 SRC) produced by crossing a plant of CMS K346 as a female with pollen of K346 SRC BC₆F₁to produce male-sterile plants heterozygous for all three mutations. Progeny plants of the CMS K346×K346 SRC BC₆F₁cross are male sterile. A plurality of CMS K346×K346 SRC BC₆F₁CMS plants (e.g., CMS F₁progeny plants) are screened for the cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S mutations to identify plants heterozygous for all three mutations. A single male-sterile plant resulting from this cross is backcrossed as a female to K346 SRC to prepare BC₇F₁CMS progeny. BC₇F₁CMS progeny homozygous for the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S mutations are identified by genotyping and designated as CMS K346 SRC. Because the CMS K346 SRC line is male sterile, it is maintained via pollination with K346 SRC. K346 SRC is crossed as the male parent to CMS K346 SRC to prepare CMS K346 SRC F₁progeny plants.

CMS K346 SRC and CMS K346 SRC F₁progeny plants have genetic backgrounds that are at least 95%, at least 97%, at least 98%, or at least 99% similar to K346. CMS K346 SRC and CMS K346 SRC F₁progeny plants exhibit low nornicotine levels and produce leaves with reduced potential for accumulating derived NNN during curing, storage, and smoking.

NC1562-1 SRC

The present disclosure also provides tobacco cultivars, and parts thereof, from NC1562-1 SRC, where representative sample seeds of this cultivar have been deposited with the ATCC under ATCC Accession No. PTA-13457. The present disclosure also includes a tobacco plant, or part thereof, produced by growing a seed of NC 1562-1 SRC. A plant of the present disclosure can include a plant with all, or essentially all, of the morphological and physiological characteristics of cultivar NC1562-1 SRC. While not being limited by process, NC1562-1 SRC is a result of introducing the cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S mutations from DH98-325-6. F₁individuals originating from a cross between NC1562-1 and DH98-325-6 and heterozygous for each mutation are then backcrossed seven times to NC1562-1 to produce BC₇F₁progeny. BC₇F₁individuals heterozygous for all three mutations are self-pollinated to produce BC₇F₂seed and individuals homozygous for all three mutations identified. A single BC₇F₂plant is self-pollinated to produce BC₇F₃(NC1562-1 SRC) in which the wild-type CYP82E4, CYP82E5v2 and CYP82E10 alleles of K346 are replaced by the mutant (e.g., cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S).

NC1562-1 SRC has a genetic background that is at least 95%, at least 97%, at least 98%, or at least 99% similar to NC1562-1. NC1562-1 SRC exhibits low nornicotine levels and produce leaves with reduced potential for accumulating derived NNN during curing, storage, and smoking.

NCTG-61 SRC

The present disclosure also provides tobacco cultivars, and parts thereof, from NCTG-61 SRC, where representative sample seeds of this cultivar have been deposited with the ATCC under ATCC Accession No. PTA-120314. The present disclosure also includes a tobacco plant, or part thereof, produced by growing a seed of NCTG-61 SRC. A plant of the present disclosure can include a plant with all, or essentially all, of the morphological and physiological characteristics of cultivar NCTG-61 SRC. While not being limited by process, NCTG-61 SRC is a result of introducing the cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S mutations from DH98-325-6. F₁individuals originating from a cross between NCTG-61 and DH98-325-6 and heterozygous for each mutation are then backcrossed seven times to NCTG-61 to produce BC₇F₁progeny. BC₇F₁individuals heterozygous for all three mutations are self-pollinated to produce BC₇F₂seed and individuals homozygous for all three mutations identified. A single BC₇F₂plant is self-pollinated to produce BC₇F₃(NCTG-61 SRC) in which the wild-type CYP82E4, CYP82E5v2 and CYP82E10 alleles of K346 are replaced by the mutant (e.g., cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S).

NCTG-61 SRC has a genetic background that is at least 95%, at least 97%, at least 98%, or at least 99% similar to NCTG-61. NCTG-61 SRC exhibits low nornicotine levels and produce leaves with reduced potential for accumulating derived NNN during curing, storage, and smoking.

CMS NCTG-61 SRC

The present disclosure also provides tobacco cultivars, and parts thereof, from CMS NCTG-61 SRC, where representative sample seeds of this cultivar have been deposited with the ATCC under ATCC Accession No. PTA-13456. The present disclosure also includes a tobacco plant, or part thereof, produced by growing a seed of CMS NCTG-61 SRC. A plant of the present disclosure can include a plant with all, or essentially all, of the morphological and physiological characteristics of cultivar CMS NCTG-61 SRC.

CMS NCTG-61 SRC is a male-sterile (CMS) version of NCTG-61 (CMS NCTG-61) produced by crossing a plant of CMS NCTG-61 as a female with pollen of NCTG-61 SRC to produce male-sterile plants heterozygous for all three mutations. The CMS progeny plants of the CMS NCTG-61×NCTG-61 SRC BC₆F₁cross are male sterile. A plurality of CMS NCTG-61×NCTG-61 SRC BC₆F₁CMS plants (e.g., CMS F₁progeny plants) are screened for the cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S mutations to identify plants heterozygous for all three mutations. A single male-sterile plant resulting from this cross is backcrossed as a female to NCTG-61 SRC to prepare BC₇F₁CMS progeny. BC₇F₁CMS progeny homozygous for the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S mutations are identified by genotyping and designated as CMS NCTG-61 SRC. Because the CMS NCTG-61 SRC line is male sterile, it is maintained via pollination with NCTG-61 SRC. NCTG-61 SRC is crossed as the male parent to CMS NCTG-61 SRC to prepare CMS NCTG-61 SRC F₁progeny plants.

CMS NCTG-61 SRC and CMS NCTG-61 SRC F₁progeny plants have genetic backgrounds that are at least 95%, at least 97%, at least 98%, or at least 99% similar to NCTG-61. CMS NCTG-61 SRC and CMS NCTG-61 SRC F₁progeny plants exhibit low nornicotine levels and produce leaves with reduced potential for accumulating derived NNN during curing, storage, and smoking.

NC196 SRC

The present disclosure includes tobacco cultivars, and parts thereof, from NC196 SRC. In another aspect, the present disclosure includes a tobacco plant, or part thereof, produced by growing the seed of NC196 SRC. A plant of the present disclosure can include a plant with all, or essentially all, of the morphological and physiological characteristics of cultivar NC196 SRC.

While not being limited by process, NC 196 SRC is produced by pollinating plants of CMS NCTG-61 SRC with pollen of NC1562-1 SRC. Again, not limited by any particular scientific theory, cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S mutations all encode for proteins with reduced or eliminated ability to convert nicotine or nornicotine. NC196 SRC has a genetic background that is at least 95%, at least 97%, at least 98%, or at least 99% similar to flue-cured tobacco cultivar NC 196, a hybrid generated by pollinating plants of a male-sterile breeding line NCTG-61 with pollen produced by fertile breeding line NC1562-1. NC 196 SRC exhibits low NNN and is not subject to conversion to high NNN's.

Other Plants

The present disclosure includes a tobacco seed produced by crossing two parent tobacco plants and harvesting the resultant tobacco seed, where at least one parent tobacco plant is K326 SRC. In one aspect, the K326 SRC is the male parent plant. In another aspect, the CMS K326 SRC is the female parent plant. One aspect of the present disclosure provides tobacco plants that are homozygous at the cyp82e4, cyp82e5v2, and cyp82E10 loci for SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 11, respectively, and which share a genetic background that is greater than 75%, 80%, 85%, 90%, 95%, 98%, or 99% similar to K326 or CMS K326. In one aspect, approximately or greater than 50%, 75%, or 100% of a progeny's genetics is provided by a plant of the present disclosure that is homozygous at the cyp82e4, cyp82e5v2, and cyp82E10 loci for SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 11, respectively. In one aspect, a plant of the present disclosure has a genetic background that is at least 95%, at least 97%, at least 98%, or at least 99% similar to K326 or CMS K326. In another aspect, a plant of the present disclosure exhibits low nornicotine and is not subject to conversion to high NNN's. In one aspect, a plant of the present disclosure is the progeny plant of a female or male parent plant that is Fusarium wilt resistant. In another aspect, a plant of K326 SRC has low resistance to black shank and moderate resistance to bacterial wilt.

The present disclosure includes a tobacco seed produced by crossing two parent tobacco plants and harvesting the resultant tobacco seed, where at least one parent tobacco plant is K346 SRC. In one aspect, the K346 SRC is the male parent plant. In another aspect, the CMS K346 SRC is the female parent plant. One aspect of the present disclosure provides tobacco plants that are homozygous at the cyp82e4, cyp82e5v2, and cyp82E10 loci for SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 11, respectively, and which share a genetic background that is greater than 75%, 80%, 85%, 90%, 95%, 98%, or 99% similar to K346 or CMS K346. In one aspect, approximately or greater than 50%, 75%, or 100% of a progeny's genetics is provided by a plant of the present disclosure that is homozygous at the cyp82e4, cyp82e5v2, and cyp82E10 loci for SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 11, respectively. In one aspect, a plant of the present disclosure has a genetic background that is at least 95%, at least 97%, at least 98%, or at least 99% similar to K346 or CMS K346. In another aspect, a plant of the present disclosure exhibits low nornicotine and is not subject to conversion to high NNN's. In one aspect, a plant of the present disclosure is the progeny plant of a female or male parent plant that is Fusarium wilt resistant. In another aspect, a plant of K346 SRC has high resistance to black shank and high resistance to bacterial wilt.

The present disclosure includes a tobacco seed produced by crossing two parent tobacco plants and harvesting the resultant tobacco seed, where at least one parent tobacco plant is NC1562-1 SRC. In one aspect, the NC1562-1 SRC is the male parent plant. In another aspect, the CMS NCTG-61 SRC is the female parent plant. One aspect of the present disclosure provides tobacco plants that are homozygous at the cyp82e4, cyp82e5v2, and cyp82E10 loci for SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 11, respectively, and which share a genetic background that is greater than 75%, 80%, 85%, 90%, 95%, 98%, or 99% similar to NC1562-1 or CMS NC1562-1. In one aspect, approximately or greater than 50%, 75%, or 100% of a progeny's genetics is provided by a plant of the present disclosure that is homozygous at the cyp82e4, cyp82e5v2, and cyp82E10 loci for SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 11, respectively. In one aspect, a plant of the present disclosure has a genetic background that is at least 95%, at least 97%, at least 98%, or at least 99% similar to NC1562-1 SRC and CMS NCTG-61 SRC. In another aspect, a plant of the present disclosure exhibits low nornicotine and is not subject to conversion to high NNN's. In one aspect, a plant of the present disclosure is the progeny plant of a female or male parent plant that is Fusarium wilt resistant. In another aspect, a plant of NC196 SRC has moderate resistance to black shank and moderate resistance to bacterial wilt.

In one aspect, a plant of the present disclosure is a medium-late maturing variety with moderately high yield potential. In another aspect, a plant of the present disclosure offers a broad range of important agronomic characteristics. In a further aspect, a plant of the present disclosure has one, two, three, four or more of the traits including, but not limited to, moderate resistance to black shank, some tolerance to blue mold, black root rot resistance, and resistance to common virus diseases. In another aspect, a plant of the present disclosure has blue mold tolerance and level 4 resistance to both races of black shank and high root rot resistance. In one aspect, a plant of the present disclosure, such as K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC, lacks Fusarium wilt resistance. In another aspect, a plant of the present disclosure is Fusarium wilt resistant. In another aspect, a plant of the present disclosure has low resistance to black shank and moderate resistance to bacterial wilt.

In an aspect, the plants of the present disclosure have reduced or eliminated ability to convert nicotine to nornicotine. In an aspect, the percentage nicotine conversion is less than 75%, 70%, 60%, 50%, or 25% of that found in K326, K346, NC1562-1, NCTG-61 or hybrid cultivar NC196. The nicotine conversion in plants of the present disclosure, including, but not limited to, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC, can be less than about 4%, about 3.5%, about 3%, about 2.5%, about 2%, about 1.5%, about 1%, about 3-1%, about 3-0.5%, or about 2-0.5%. In a preferred aspect, the percentage nicotine conversion is less than 25%, 10%, 5%, or 2% of that found in K326, K346, NC1562-1, NCTG-61 or hybrid cultivar NC196 without the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S mutations. In an aspect, the tobacco plants of the present disclosure have a nicotine conversion rate of 3.5, 3.25, 3.0 or 2.75% or less. In another aspect, the nicotine conversion rate of tobacco plants of the present disclosure can be 4.0, 3.9, 3.8, 3.7, 3.6, 3.5, 3.4, 3.3, 3.2, 3.1, 3.0, 2.9, 2.8, 2.7, 2.6, 2.5, 2.4, 2.3, 2.2, 2.1, 2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5% or less. In another aspect, the nicotine conversion rate of tobacco plants of the present disclosure can be 2.9, 2.8, 2.7, 2.6, 2.5, 2.4, 2.3, 2.2, 2.1, 2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6% or less. In another aspect, the nicotine conversion rates can be from 0.5 to 0.9%, 0.5 to 1.5%, 0.5 to 2.0%, 0.5 to 2.5%, 0.5 to 2.75%, and 0.5 to 3.0%. In another aspect, the nicotine conversion rates can be from 1.0 to 1.5%, 1.0 to 1.75%, 1.0 to 2.0%, 1.0 to 2.5%, 1.0 to 2.75%, and 1.0 to 3.0%. In another aspect, the nicotine conversion rate in a plant of the present disclosure may be less than 2.9, 2.75, 2.5, 2.25, 2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1 or 1.0%. In an aspect, the tobacco plants of the present disclosure have a nicotine conversion rate of 3.5, 3.25, 3.0 or 2.75% or less. In another aspect, the nicotine conversion rate of tobacco plants of the present disclosure can be 2.9, 2.8, 2.7, 2.6, 2.5, 2.4, 2.3, 2.2, 2.1, 2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6% or less. In another aspect, the nicotine conversion rates can be from 0.5 to 0.9%, 0.5 to 1.5%, 0.5 to 2.0%, 0.5 to 2.5%, 0.5 to 2.75%, and 0.5 to 3.0%. In another aspect, the nicotine conversion rates can be from 1.0 to 1.5%, 1.0 to 1.75%, 1.0 to 2.0%, 1.0 to 2.5%, 1.0 to 2.75%, and 1.0 to 3.0%. In another aspect, the nicotine conversion rate in a plant of the present disclosure may be less than 2.9, 2.75, 2.5, 2.25, 2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1 or 1.0%.

In a further aspect, tobacco filler made from a plant of the present disclosure comprise a nicotine conversion rate between 0.05 and 0.5%, between 0.1 and 0.45%, between 0.15 and 0.4%, between 0.2 and 0.35%, or between 0.25 and 0.3%.

In a further aspect, tobacco filler made from a plant of the present disclosure comprise a nicotine conversion rate between 0.05 and 0.5%, between 0.1 and 0.5%, between 0.15 and 0.5%, between 0.2 and 0.5%, between 0.25 and 0.5%, between 0.3 and 0.5%, between 0.35 and 0.5%, between 0.4 and 0.5%, or between 0.45 and 0.5%.

In a further aspect, tobacco filler made from a plant of the present disclosure comprise a nicotine conversion rate between 0.05 and 0.5%, between 0.05 and 0.45%, between 0.05 and 0.4%, between 0.05 and 0.35%, between 0.05 and 0.3%, between 0.05 and 0.25%, between 0.05 and 0.2%, between 0.05 and 0.15%, or between 0.05 and 0.1%.

In another aspect, the tobacco plants of the present disclosure typically have a reduced amount of nornicotine of less than about 0.10% dry weight. For example, the nornicotine content in such plants can be 1.2, 1.0, 0.7, 0.5, 0.4, 0.2, 0.1, 0.09, 0.085, 0.08, 0.075, 0.07, 0.065, 0.06, 0.055, 0.05, 0.045, 0.04, 0.035, 0.025, 0.01, 0.009, 0.0075, 0.005, 0.0025, 0.001, 0.0009, 0.00075, 0.0005, 0.00025, or 0.0001% dry weight, or undetectable. In another aspect, the nornicotine content can be less than 1.2, 1.0, 0.9, 0.8, 0.7, 0.5, 0.4, 0.2, 0.1, 0.075, 0.05, 0.025, 0.01, 0.009, 0.0075, 0.005, 0.0025, 0.001, 0.0009, 0.00075, 0.0005, 0.00025, or 0.0001% dry weight. In another aspect, the nornicotine content in such plants can be from 1.2-1.0, 0.7-0.5, 0.4-0.2, 0.1-0.075, 0.05-0.025, 0.01-0.0075, 0.005-0.0025, 0.001-0.00075, 0.0005-0.00025, or 0.0005-0.0001% dry weight. In a plant of the present disclosure, the nornicotine is a relatively small percentage of total alkaloids in the plant compared to a commercial seedlot of K326, K346, NC1562-1, NCTG-61 and hybrid cultivar NC196. The nornicotine in a plant of the present disclosure may be 2-1%, less than 3%, about 2%, about 1.5%, about 1%, or 0.75% of total alkaloids. Tobacco products having a reduced amount of nitrosamine content can be manufactured using tobacco plant material from plants and plant parts of the present disclosure. The tobacco product typically has a reduced amount of nornicotine of less than about 3 mg/g. For example, the nornicotine content in such a product can be 3.0 mg/g, 2.5 mg/g, 2.0 mg/g, 1.5 mg/g, 1.0 mg/g, 750 μg/g, 500 pg/g, 250 pg/g, 100 pg/g, 75 pg/g, 50 pg/g, 25 pg/g, 10 pg/g, 7.0 pg/g, 5.0 pg/g, 4.0 pg/g, 2.0 pg/g, 1.0 pg/g, 0.5 pg/g, 0.4 pg/g, 0.2 pg/g, 0.1 pg/g, 0.05 pg/g, 0.01 pg/g, or undetectable. The tobacco product typically has a reduced amount of NNN of less than about 10 pg/g. For example, the nornicotine content in such a product can be about 10 pg/g, 7.0 pg/g, 5.0 pg/g, 4.0 pg/g, 2.0 pg/g, 1.0 pg/g, 0.5 pg/g, 0.4 pg/g, 0.2 pg/g, 0.1 pg/g, 0.05 pg/g, 0.01 pg/g, or undetectable. The percentage of secondary alkaloids relative to total alkaloid content contained in a plant of the present disclosure may not be statistically different than from a commercial seedlot of K326, K346, NC1562-1, NCTG-61 and hybrid cultivar NC196.

In an aspect, a plant of the present disclosure or tobacco filler made therefrom has reduced level of NNN. In an aspect, the NNN level is less than 75%, 70%, 60%, 50%, or 25% of that found in K326, K346, NC1562-1, NCTG-61 or hybrid cultivar NC196. The NNN level in tobacco filler made from plants of the present disclosure, including, but not limited to, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC, can be less than about 0.5, 0.4, 0.3, 0.2, or 0.1 ppm. In a preferred aspect, the NNN level of a tobacco filler disclosed herein is between 0.8 and 0.05, between 0.8 and 0.1, between 0.8 and 0.2, between 0.8 and 0.3, between 0.8 and 0.4, between 0.8 and 0.5, between 0.8 and 0.6, between 0.7 and 0.05, between 0.6 and 0.05, between 0.5 and 0.05, between 0.4 and 0.05, between 0.3 and 0.05, between 0.2 and 0.05, between 0.1 and 0.05, between 0.7 and 0.1, between 0.6 and 0.2, or between 0.5 and 0.3 ppm. In a preferred aspect, the NNN level is less than 25%, 10%, 5%, or 2% of that found in K326, K346, NC1562-1, NCTG-61 or hybrid cultivar NC196 without the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S mutations.

In an aspect, a cigarette made from a plant of the present disclosure has reduced level of NNN in its smoke. In an aspect, the cigarette smoke NNN level from plants of the instant disclosure is less than 75%, 70%, 60%, 50%, or 25% of that found in K326, K346, NC1562-1, NCTG-61 or hybrid cultivar NC196. The cigarette smoke NNN level from plants of the present disclosure, including, but not limited to, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC, can be less than about 0.035, 0.03, 0.025, 0.02, 0.015, 0.01, or 0.005 ng/mg TPM. In a preferred aspect, the cigarette smoke NNN level is between 0.035 and 0.005, between 0.035 and 0.01, between 0.035 and 0.015, between 0.035 and 0.02, between 0.035 and 0.025, between 0.03 and 0.005, between 0.025 and 0.005, between 0.02 and 0.005, between 0.015 and 0.005, between 0.03 and 0.01, or between 0.025 and 0.015 ng/mg TPM. In a preferred aspect, the cigarette smoke NNN level is less than 25%, 10%, 5%, or 2% of that found in K326, K346, NC1562-1, NCTG-61 or hybrid cultivar NC196 without the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S mutations.

Differences between two inbred tobacco varieties or two hybrid tobacco varieties can be evaluated using statistical approaches. Statistical analysis includes the calculation of mean values, determination of the statistical significance of the sources of variation, and the calculation of the appropriate variance components. Methods for determining statistical significance are known in the art. Statistical software is available, for example, the PROC GLM function of SAS. Significance is generally presented as a “p-value.” A statistically significant p-value is less than 0.10. In a preferred aspect, the p-value is less than, or equal to, 0.05. In another aspect, the p-value is 0.04 or less, 0.03 or less, or 0.02 or less. In yet another aspect, a statistically significant value is less than 0.01. In yet another aspect, it can be less than 0.009, less than 0.008, less than 0.007, less than 0.006, less than 0.005, less than 0.004, less than 0.003, less than 0.002, or less than 0.001.

Tobacco plants of the present disclosure that are homozygous for the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S alleles have a reversion rate that is statistically significantly lower than corresponding control low-converter plants having wild type nicotine demethylase CYP82E4, E5, and E10 genes. In addition, homozygous CYP82E4, CYP82E5, and CYP82E10 triple mutant tobacco plants have a percent conversion to nornicotine of less than 2.0%, e.g., undetectable to 2.0%, 1.0 to 2.0%, 0.8 to 1.8%, 0.8 to 2.0%, or 1.0 to 2.0%.

Nicotine and nornicotine can be measured in ethylene-treated leaves using methods known in the art (e.g., gas chromatography). Percent nicotine demethylation in a sample is calculated by dividing the level of nornicotine by the combined level of nicotine and nornicotine as measured in the sample, and multiplying by 100. Percent nicotine demethylation in a sample from a plant of the present disclosure is 50, 40, 30, 20, or 10 percent of a sample from an individual plant grown from a commercial seedlot of K326, K346, NC1562-1, NCTG-61 and hybrid cultivar NC196.

In an aspect, the tobacco plants of the present disclosure have a USDA quality index of about 73, about 72, about 71, about 70, about 69, about 68, about 67 or about 66. In an aspect, the tobacco plants of the present disclosure have a USDA quality index of about 65. In another aspect, the quality index may be at least about 55, 60, 62.5 or greater. In another aspect, tobacco plants of the present disclosure can have a quality index of 60-65, 60-70, 62.5-65, 62.5-70, or 65-70.

A plant of the present disclosure, including, but not limited to, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC, can have any yield, including high (e.g., over 3000 lbs/A), moderately high (e.g., 2200-3000 lbs/A), and moderate (e.g., less than 2000 lbs/A) yield potential.

In another aspect, the present disclosure also provides for a plant grown from the seed of a K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC plant in which alkaloids obtained from tobacco plants grown for the seed have decreased nornicotine, as well as plant parts and tissue cultures from such plants, representative sample seeds of these cultivars having been deposited with the ATCC, for example, under ATCC Accession No. PTA-13455 for K326 SRC, ATCC Accession No. PTA-13454 for CMS K326 SRC, ATCC Accession No. PTA-13453 for K346 SRC, ATCC Accession No. PTA-13452 for CMS K346 SRC, ATCC Accession No. PTA-13457 for NC1562-1 SRC, ATCC Accession No. PTA-120314 for NCTG-61 SRC, ATCC Accession No. PTA-13456 for CMS NCTG-61 SRC, and ATCC Accession No. PTA-13458 for hybrid cultivar NC196 SRC.

An aspect of the present disclosure provides for parts of the cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC. For example, leaves, pollen, embryos, endosperm, cotyledons, hypocotyls, roots, root tips, anthers, flowers, ovules, shoots, stems, stalks, pith and capsules, tissue culture comprising tissue, callus, cells or protoplasts of the cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC. In another aspect, the present disclosure provides for parts from hybrids of cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC derived tobacco plants. In yet another aspect, the present disclosure provides for parts from genetically modified (e.g., by conventional breeding or genetic engineering techniques) forms of the foregoing plants and tissue culture.

In a further aspect, the present disclosure also provides for a non-regenerable cell of the cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC. In another aspect, the present disclosure also provides for non-reproductive material or a non-reproductive cell of the cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC.

Additional aspects of the present disclosure provide products comprising tobacco wherein the tobacco further comprises tobacco from the plants of the present disclosure, and parts thereof. Other aspects of the disclosure provide cured plant leaves and other plant parts. Accordingly, in some aspects, the cured plant parts include, but are not limited to, a leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, protoplast, root, root tip, pistil, anther, flower, shoot, stem, pod, petiole, and the like, and combinations thereof.

Thus, in some aspects, the present disclosure provides a cured tobacco comprising the leaves of the tobacco plant designated K326 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455. In another aspect, the present disclosure provides a cured tobacco comprising the leaves of the tobacco plant designated CMS K326 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454. In another aspect, the present disclosure provides a cured tobacco comprising the leaves of the tobacco plant designated K346 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453. In another aspect, the present disclosure provides a cured tobacco comprising the leaves of the tobacco plant designated CMS K346 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452. In a further aspect, the present disclosure provides a cured tobacco comprising the leaves of the tobacco plant designated NC1562-1 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13457. In yet another aspect, the present disclosure provides a cured tobacco comprising the leaves of the tobacco plant designated NCTG-61 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314. In yet another aspect, the present disclosure provides a cured tobacco comprising the leaves of the tobacco plant designated CMS NCTG-61 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456. In yet another aspect, the present disclosure provides a cured tobacco comprising the leaves of the hybrid tobacco plant designated NC 196 SRC, a representative sample seed of said hybrid cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458.

In an aspect, the present disclosure provides a cured tobacco comprising the stems of the tobacco plant designated K326 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455. In another aspect, the present disclosure provides a cured tobacco comprising the stems of the tobacco plant designated CMS K326 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454. In an aspect, the present disclosure provides a cured tobacco comprising the stems of the tobacco plant designated K346 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453. In another aspect, the present disclosure provides a cured tobacco comprising the stems of the tobacco plant designated CMS K346 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452. In a further aspect, the present disclosure provides a cured tobacco comprising the stems of the tobacco plant designated NC1562-1 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13457. In yet another aspect, the present disclosure provides a cured tobacco comprising the stems of the tobacco plant designated NCTG-61 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314. In yet another aspect, the present disclosure provides a cured tobacco comprising the stems of the tobacco plant designated CMS NCTG-61 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456. In yet another aspect, the present disclosure provides a cured tobacco comprising the stems of the hybrid tobacco plant designated NC 196 SRC, a representative sample seed of said hybrid cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13458.

In an aspect, the present disclosure provides a cured tobacco comprising the leaves and stems of the tobacco plants designated K326 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13455. In another aspect, the present disclosure provides a cured tobacco comprising the leaves and stems of the tobacco plants designated CMS K326 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13454. In an aspect, the present disclosure provides a cured tobacco comprising the leaves and stems of the tobacco plants designated K346 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13453. In another aspect, the present disclosure provides a cured tobacco comprising the leaves and sterns of the tobacco plants designated CMS K346 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13452. In a further aspect, the present disclosure provides a cured tobacco comprising the leaves and stems of the tobacco plants designated NC1562-1 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No, PTA-13457. In yet another aspect, the present disclosure provides a cured tobacco comprising the leaves and stems of the tobacco plants designated NCTG-61 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-120314. In yet another aspect, the present disclosure provides a cured tobacco comprising the leaves and stems of the tobacco plants designated CMS NCTG-61 SRC, a representative sample seed of said cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13456. In yet another aspect, the present disclosure provides a cured tobacco comprising the leaves and stems of the hybrid tobacco plants designated NC196 SRC, a representative sample seed of said hybrid cultivar having been deposited with the ATCC under ATCC Accession No. PTA-13458.

The present disclosure also provides a container of K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC seeds or other seeds of the present disclosure in which alkaloids obtained from tobacco plants grown from greater than 50% of the seeds have decreased nornicotine. In another aspect, alkaloids obtained from K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC plants or other plants of the present disclosure grown from greater than 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the seeds in the container have decreased nornicotine, representative samples of seeds of these cultivars having been deposited with the ATCC, for example, under ATCC Accession No. PTA-13455 for K326 SRC, ATCC Accession No. PTA-13454 for CMS K326 SRC, ATCC Accession No. PTA-13453 for K346 SRC, ATCC Accession No. PTA-13452 for CMS K346 SRC, ATCC Accession No. PTA-13457 for NC1562-1 SRC, ATCC Accession No. PTA-120314 for NCTG-61 SRC, ATCC Accession No. PTA-13456 for CMS NCTG-61 SRC, and ATCC Accession No. PTA-13458 for hybrid cultivar NC196 SRC.

The container of K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC seeds or other seeds of the present disclosure may contain any number, weight or volume of seeds. For example, a container can contain at least, or greater than, about 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1500, 2000, 2500, 3000, 3500, 4000 or more seeds. Alternatively, the container can contain at least, or greater than, about 1 ounce, 5 ounces, 10 ounces, 1 pound, 2 pounds, 3 pounds, 4 pounds, 5 pounds or more seeds. Representative samples of seeds of these cultivars having been deposited with the ATCC, for example, under ATCC Accession No. PTA-13455 for K326 SRC, ATCC Accession No. PTA-13454 for CMS K326 SRC, ATCC Accession No. PTA-13453 for K346 SRC, ATCC Accession No. PTA-13452 for CMS K346 SRC, ATCC Accession No. PTA-13457 for NC1562-1 SRC, ATCC Accession No. PTA-120314 for NCTG-61 SRC, ATCC Accession No. PTA-13456 for CMS NCTG-61 SRC, and ATCC Accession No. PTA-13458 for hybrid cultivar NC196 SRC.

Containers of K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC seeds or other seeds of the present disclosure may be any container available in the art. By way of a non-limiting example, a container may be a box, a bag, a packet, a pouch, a tape roll, a pail, a foil, a tube, or a bottle. Representative samples of seeds of these cultivars having been deposited with the ATCC, for example, under ATCC Accession No. PTA-13455 for K326 SRC, ATCC Accession No, PTA-13454 for CMS K326 SRC, ATCC Accession No. PTA-13453 for K346 SRC, ATCC Accession No. PTA-13452 for CMS K346 SRC, ATCC Accession No. PTA-13457 for NC1562-1 SRC, ATCC Accession No. PTA-120314 for NCTG-61 SRC, ATCC Accession No, PTA-13456 for CMS NCTG-61 SRC, and ATCC Accession No. PTA-13458 for hybrid cultivar NC196 SRC.

In another aspect, the present disclosure also provides a container of K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid cultivar NC196 SRC in which greater than 50% of K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC seeds or other seeds of the present disclosure have decreased nornicotine. Representative samples of seeds of these cultivars having been deposited with the ATCC, for example under ATCC Accession No. PTA-13455 for K326 SRC, ATCC Accession No. PTA-13454 for CMS K326 SRC, ATCC Accession No. PTA-13453 for K346 SRC, ATCC Accession No. PTA-13452 for CMS K346 SRC, ATCC Accession No. PTA-13457 for NC1562-1 SRC, ATCC Accession No. PTA-120314 for NCTG-61 SRC, ATCC Accession No. PTA-13456 for CMS NCTG-61 SRC, and ATCC Accession No. PTA-13458 for hybrid cultivar NC196 SRC.

In one aspect, the present disclosure provides a seed of a K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC plant or other plant of the present disclosure in which a plant grown from a seed is male sterile. Representative samples of seeds of these cultivars having been deposited with the ATCC, for example, under ATCC Accession No. PTA-13455 for K326 SRC, ATCC Accession No. PTA-13454 for CMS K326 SRC, ATCC Accession No. PTA-13453 for K346 SRC, ATCC Accession No. PTA-13452 for CMS K346 SRC, ATCC Accession No. PTA-13457 for NC1562-1 SRC, ATCC Accession No. PTA-120314 for NCTG-61 SRC, ATCC Accession No. PTA-13456 for CMS NCTG-61 SRC, and ATCC Accession No. PTA-13458 for hybrid cultivar NC196 SRC.

The instant disclosure further provides cured tobacco material, tobacco blends, and tobacco products made from tobacco plants disclosed herein. In one aspect, the cured tobacco material of the instant disclosure is flue-cured. In another aspect, the cured tobacco material of the instant disclosure is sun-cured, air-cured, or fire-cured.

Tobacco material obtained from the tobacco lines, varieties or hybrids of the present disclosure can be used to make tobacco products including, without limitation, cigarette products (e.g., cigarettes and bidi cigarettes), cigar products (e.g., cigar wrapping tobacco and cigarillos), pipe tobacco products, products derived from tobacco, tobacco derived nicotine products, smokeless tobacco products (e.g., moist snuff, dry snuff, and chewing tobacco), films, chewables, tabs, shaped parts, gels, consumable units, insoluble matrices, hollow shapes and the like. See, e.g., U.S. Patent Publication No. US 2006/0191548, which is herein incorporated by reference in its entirety.

Tobacco products derived from plants of the present disclosure also include cigarettes and other smoking articles, particularly those smoking articles including filter elements, wherein the rod of smokeable material includes cured tobacco within a tobacco blend. In an aspect, a tobacco product may be pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, shredded tobacco, and cut tobacco.

In an aspect, a tobacco product of the instant disclosure is selected from the group consisting of a cigarillo, a non-ventilated recess filter cigarette, a vented recess filter cigarette, a cigar, snuff, pipe tobacco, cigar tobacco, cigarette tobacco, chewing tobacco, leaf tobacco, hookah tobacco, shredded tobacco, and cut tobacco. In another aspect, a tobacco product of the instant disclosure is a smokeless tobacco product. In a further aspect, a tobacco product of the instant disclosure is selected from the group consisting of loose leaf chewing tobacco, plug chewing tobacco, moist snuff, and nasal snuff. In yet another aspect, a tobacco product of the instant disclosure is selected from the group consisting of an electronically heated cigarette, an e-cigarette, an electronic vaporing device.

In an aspect, the tobacco product of the present disclosure can be a blended tobacco product. In other aspects of the disclosure, the tobacco product of the present disclosure can be a reduced nicotine tobacco product. In still other aspects, the tobacco product of the present disclosure can be a blended tobacco product with reduced nicotine content. Thus, the tobacco product of the present disclosure can be a blended reduced nicotine tobacco product. Tobacco product material comprises a blend of tobacco materials from the present disclosure, wherein the blend comprises at least about 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 95 percent by weight of a cured tobacco, based on the dry weight of the tobacco material. US 2008/0245377 is herein incorporated by reference for blend mixtures in its entirety.

In an aspect, tobacco products having a reduced amount of nitrosamine content can be manufactured using tobacco plant material from plants and plant parts of the present disclosure. The tobacco product typically has a reduced amount of nornicotine of less than about 3 mg/g. For example, the nornicotine content in such a product can be 3.0 mg/g, 2.5 mg/g, 2.0 mg/g, 1.5 mg/g, 1.0 mg/g, 750 μg/g, 500 pg/g, 250 pg/g, 100 pg/g, 75 pg/g, 50 pg/g, 25 pg/g, 10 pg/g, 7.0 pg/g, 5.0 pg/g, 4.0 pg/g, 2.0 pg/g, 1.0 pg/g, 0.5 pg/g, 0.4 pg/g, 0.2 pg/g, 0.1 pg/g, 0.05 pg/g, 0.01 pg/g, or undetectable. The tobacco product typically has a reduced amount of NNN of less than about 10 pg/g. For example, the nornicotine content in such a product can be about 10 pg/g, 7.0 pg/g, 5.0 pg/g, 4.0 pg/g, 2.0 pg/g, 1.0 pg/g, 0.5 pg/g, 0.4 pg/g, 0.2 pg/g, 0.1 pg/g, 0.05 pg/g, 0.01 pg/g, or undetectable. The percentage of secondary alkaloids relative to total alkaloid content contained in a plant of the present disclosure may not be statistically different than from a commercial seedlot of K326, K346, NC1562-1, NCTG-61 and hybrid cultivar NC196.

The instant disclosure further provides a method manufacturing a tobacco product, where the method comprising conditioning aged tobacco material made from an tobacco plant disclosed herein to increase its moisture content from between about 12.5% and about 13.5% to about 21%, blending the conditioned tobacco material to produce a desirable blend. In one aspect, the method of manufacturing a tobacco product disclosed herein further comprises casing or flavoring the blend. Generally, during the casing process, casing or sauce materials are added to blends to enhance their quality by balancing the chemical composition and to develop certain desired flavor characteristics. Further details for the casing process can be found in Tobacco Production, Chemistry and Technology, Edited by L. Davis and M. Nielsen, Blackwell Science, 1999.

A tobacco plant of the present disclosure designated K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid cultivar NC196 SRC, carrying the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S alleles can be used in a plant breeding program to create useful lines, cultivars, varieties, progeny, inbreds, and hybrids. Thus, in some aspects, an F₁, F₂, F₃, or later generation tobacco plant containing the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S alleles is crossed with a second Nicotiana plant, and progeny of the cross are identified in which the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S alleles are present. It will be appreciated that the second Nicotiana plant will be K326, K346, NC1562-1, NCTG-61 and hybrid cultivar NC196 or any other Nicotiana species or line, optionally with an additional desirable trait, such as herbicide resistance mentioned below.

In still other aspects, methods of the present disclosure further include self-pollinating or pollinating a male sterile pollen acceptor with a pollen donor capable of being used in production of a progeny of the present disclosure, such as a male sterile hybrid of the present disclosure. Either the male sterile pollen acceptor plant or the pollen donor plant has at least one mutant allele, two, or even three mutant alleles at a nicotine demethylase locus, such as the cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S. In an aspect, all three alleles at each nicotine demethylase locus are mutant alleles, making the plant homozygous for cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S.

Breeding can be carried out via any known procedures. DNA fingerprinting, SNP or similar technologies may be used in a marker-assisted selection (MAS) breeding program to transfer or breed mutant alleles of a nicotine demethylase gene into other tobaccos. For example, a breeder can create segregating populations from hybridizations of a genotype containing cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S alleles with an agronomically desirable genotype. Plants in the F₂or backcross generations can be screened using a marker developed from cyp82e4 W329Stop, the cyp82e5v2 W422Stop, or cyp82e10 P381S alleles or a fragment thereof, using one of the techniques known in the art or listed herein. Plants identified as possessing one or more cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S alleles can be backcrossed or self-pollinated to create a second population to be screened. Depending on the expected inheritance pattern or the MAS technology used, it may be necessary to self-pollinate the selected plants before each cycle of backcrossing to aid identification of the desired individual plants. Backcrossing or other breeding procedure can be repeated until the desired phenotype of the recurrent parent is recovered. A recurrent parent in the present disclosure can be K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or NC 196 SRC. Other breeding techniques can be found, for example, in Wernsman, E. A., and Rufty, R. C. 1987. Chapter Seventeen. Tobacco. Pages 669-698 In: Cultivar Development. Crop Species. W. H. Fehr (ed.), MacMillan Publishing Go., Inc., New York, N.Y., incorporated herein by reference in their entirety.

Nicotiana species which exhibit breeding compatibility with Nicotiana tabacum include Nicotiana amplexicaulis, PI 271989; Nicotiana benthamiana PI 555478; Nicotiana bigelovii PI 555485; Nicotiana debneyi; Nicotiana excelsior PI 224063; Nicotiana glutinosa PI 555507; Nicotiana goodspeedii PI 241012; Nicotiana gossei PI 230953; Nicotiana hesperis PI 271991; Nicotiana knightiana PI 555527; Nicotiana maritima PI 555535; Nicotiana megalosiphon PI 555536; Nicotiana nudicaulis PI 555540; Nicotiana paniculata PI 555545; Nicotiana plumbaginifolia PI 555548; Nicotiana repanda PI 555552; Nicotiana rustica; Nicotiana suaveolens PI 230960; Nicotiana sylvestris PI 555569; Nicotiana tomentosa PI 266379; Nicotiana tomentosiformis; and Nicotiana trigonophylla PI 555572. See also, Compendium of Tobacco Diseases published by American Phytopathology Society, or The Genus Nicotiana Illustrated, published by Japan Tobacco Inc, hereby incorporated by reference in their entirety.

The result of a plant breeding program using the mutant tobacco plants described herein includes useful lines, cultivars, varieties, progeny, inbreds, and hybrids. As used herein, the term “cultivar” or “variety” refers to a population of plants that share constant characteristics which separate them from other plants of the same species. A cultivar or variety is often, although not always, sold commercially. While possessing one or more distinctive traits, a cultivar or variety is further characterized by a very small overall variation between individuals within that variety. A “pure line” variety may be created by several generations of self-pollination and selection, or vegetative propagation from a single parent using tissue or cell culture techniques. A cultivar or variety can be essentially derived from another cultivar, line, or variety. As defined by the International Convention for the Protection of New Varieties of Plants (Dec. 2, 1961, as revised at Geneva on Nov. 10, 1972, on Oct. 23, 1978, and on Mar. 19, 1991), a cultivar or variety is “essentially derived” from an initial cultivar or variety if: a) it is predominantly derived from the initial cultivar or variety, or from a cultivar or variety that is predominantly derived from the initial cultivar or variety, while retaining the expression of the essential characteristics that result from the genotype or combination of genotypes of the initial cultivar or variety; b) it is clearly distinguishable from the initial cultivar or variety; and c) except for the differences which result from the act of derivation, it conforms to the initial cultivar or variety in the expression of the essential characteristics that result from the genotype or combination of genotypes of the initial cultivar or variety. Essentially derived varieties can be obtained, for example, by the selection of a natural or induced mutant, a somaclonal variant, a variant individual from plants of the initial variety, backerossing, or transformation. A “line” as distinguished from a cultivar or variety most often denotes a group of plants used non-commercially, for example in plant research. A line typically displays little overall variation between individuals for one or more traits of interest, although there may be some variation between individuals for other traits.

Hybrid tobacco varieties can be produced by preventing self-pollination of female parent plants (i.e., seed parents) of a first variety, permitting pollen from male parent plants of a second variety to fertilize the female parent plants, and allowing F₁hybrid seeds to form on the female plants. Self-pollination of female plants can be prevented by emasculating the flowers at an early stage of flower development. Alternatively, pollen formation can be prevented on the female parent plants using a form of male sterility. For example, male sterility can be produced by cytoplasmic male sterility (CMS), or transgenic male sterility wherein a transgene inhibits microsporogenesis and/or pollen formation, or self-incompatibility. Female parent plants containing CMS are particularly useful. In aspects in which the female parent plants are CMS, pollen may be harvested from male fertile plants and applied manually to the stigmas of CMS female parent plants, and the resulting F₁seed is harvested.

Plants can be used to form single-cross tobacco F₁hybrids. Alternatively, three-way crosses can be carried out wherein a single-cross F₁hybrid is used as a female parent and is crossed with a different male parent. As another alternative, double-cross hybrids can be created wherein the F₁progeny of two different single-crosses are themselves crossed. Self-incompatibility can be used to particular advantage to prevent self-pollination of female parents when forming a double-cross hybrid.

Successful crosses yield F₁plants that are fertile, have cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S alleles, and can be backcrossed with one of the parents, such as K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or NC 196 SRC, if desired. In some aspects, a plant population in the F₂generation is screened for cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S alleles. Selected plants can be crossed with one of the parents and the first backcross (BC1) generation plants are self-pollinated to produce a BC1 F₂population that is again screened for variant nicotine demethylase gene expression (e.g., the null version of the nicotine demethylase gene). The process of backcrossing, self-pollination, and screening is repeated, for example, at least 4 times, until the final screening produces a plant that is fertile and reasonably similar to the recurrent parent. This plant, if desired, is self-pollinated and the progeny are subsequently screened again to confirm that the plant exhibits the same low nicotine conversion phenotype as K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or NC 196 SRC. Breeder's seed of the selected plant is produced using standard methods including, for example, field testing, confirmation of the null condition for nicotine demethylase, chemical analyses of cured leaf to determine the level of alkaloids and/or chemical analyses of cured leaf to determine the ratio of nornicotine to nicotine+nornicotine.

In one aspect, a F₁progeny is the result of a cross between K326 SRC and CMS K326 SRC to generate F₁progeny that are male sterile. In another aspect, a F₁progeny is the result of a cross between K346 SRC and CMS K346 SRC to generate F₁progeny that are male sterile. Male sterile tobacco plants may be produced by any method known in the art. Methods of producing male sterile tobacco are described in Wernsman, E. A., and Rufty, R. C. 1987. Chapter Seventeen. Tobacco. Pages 669-698 In: Cultivar Development. Crop Species. W. H. Fehr (ed.), MacMillan Publishing Go., Inc., New York, N.Y. 761 pp.

The present disclosure further provides methods of producing a tobacco plant by crossing one of cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid NC196 SRC with itself or a different tobacco line. The disclosure further relates to methods for producing other tobacco cultivars or breeding lines derived from cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid NC196 SRC by crossing a plant of cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, CMS NCTG-61 SRC, and hybrid NC196 SRC with a second tobacco plant and growing the progeny seed to yield a K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC-derived tobacco plant. An additional aspect of the present disclosure provides a method for a tobacco plant that contains in its genetic material one or more transgenes, comprising crossing cultivars of the present disclosure with a second cultivar containing one or more transgenes wherein progeny are produced, so that the genetic material of the progeny that result from the cross comprise the transgene(s) optionally operably linked to one or more regulatory elements. In one aspect, the second cultivar may be a plant derived from cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC transformed with one or more transgenes.

The disclosure further provides for the vegetative propagation of a plant of cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid NC196 SRC, hybrids and progeny thereof. In one aspect, the disclosure provides for a method of vegetatively propagating a plant of a tobacco cultivar comprising collecting tissue capable of being propagated from a plant of cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid NC196 SRC, cultivating the tissue to obtain a proliferated shoot and rooting the proliferated shoots to obtain a rooted plantlet. In another aspect, the plant tissue may be collected from an F₁hybrid of a plant of cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid NC196 SRC. In an aspect, the plant tissue may be collected from an F₂, F₃, F₄or later progeny plant obtained by breeding a plant of cultivars K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid NC196 SRC.

A plant comprising a mutation in a nicotine demethylase gene can be identified by selecting or screening the mutagenized plant material, or progeny thereof. Such screening and selection methodologies are known to those having ordinary skill in the art. Examples of screening and selection methodologies include, but are not limited to, Southern analysis, PCR amplification for detection of a polynucleotide, Northern blots, RNase protection, primer-extension, RT-PCR amplification for detecting RNA transcripts, enzymatic assays for detecting enzyme or ribozyme activity of polypeptides and polynucleotides, and protein gel electrophoresis, Western blots, immunoprecipitation, and enzyme-linked immunoassays to detect polypeptides. Other techniques such as in situ hybridization, enzyme staining, and immunostaining also can be used to detect the presence or expression of polypeptides and/or polynucleotides. Methods for performing all of the referenced techniques are known.

It is understood that a tobacco plant of the present disclosure, including, but not limited to, K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC, and hybrid cultivar NC196 SRC, can be transformed by a genetic construct or transgene using a technique known in the art. Without limitation, an example of a desired trait is herbicide resistance, pest resistance, disease resistance, high yield, high grade index, curability, curing quality, mechanical harvestability, holding ability, leaf quality, height, plant maturation (e.g., early maturing, early to medium maturing, medium maturing, medium to late maturing, or late maturing), stalk size (e.g., small, medium, or large stalk), or leaf number per plant (e.g., small (e.g., 5-10 leaves), medium (e.g., 11-15 leaves), or large (e.g., 16-21) number of leaves), or any combination. Any plant of the present disclosure can be used as a basis for tissue culture, regenerated, transformed, or a combination of any of these. In an aspect, a plant of the present disclosure derived by tissue culture, transformation, or both has all of the morphological and physiological characteristics of cultivar K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, CMS NCTG-61 SRC, or hybrid NC196 SRC.

Having now generally described the disclosure, the same will be more readily understood through reference to the following examples that are provided by way of illustration, and are not intended to be limiting of the present disclosure, unless specified.

EXAMPLES
Example 1
Breeding of Homozygous cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S Mutant Plants into the K326 Flue-Cured Tobacco Cultivar

K326 SRC is a backcross-derived version of flue-cured tobacco cultivar K326 carrying introduced mutations in three genes (CYP82E4 (SEQ ID NO: 5), CYP82E5 (SEQ ID NO: 8), and CYP82E10 (SEQ ID NO: 9)) previously documented to encode for nicotine demethylase enzymes (Lewis et al., 2010). The introduced mutations in CYP82E4 (cyp82e4 W329Stop (SEQ ID NO: 1)) and CYP82E5 (cyp82e5v2 W422Stop (SEQ ID NO: 2)) encode for premature stop codons which render the genes non-functional. The introduced mutation in CYP82E10 (cyp82e10 P381S (SEQ ID NO: 11)) does not encode for a premature stop codon, but does render the gene product non-functional for converting nicotine to nornicotine (Lewis et al. 2010). When in homozygous condition, the three mutations result in tobacco plants with (1) reduced genetic capacity to demethylate nicotine to form nornicotine, and (2) a corresponding diminished potential to accumulate N-nitrosonornicotine (NNN), a potent carcinogen found in many tobacco products. The mutation in CYP82E4 (cyp82e4 W329Stop (SEQ ID NO: 1)) also provides phenotypic stability for the “nicotine conversion” trait and eliminates the requirement to utilize the cumbersome “LC” method for reducing levels of nornicotine in tobacco cultivars.

The original tobacco cultivar K326 is a fertile inbred line. CMS K326 is a cytoplasmic male-sterile version of K326. To develop K326 SRC, an individual plant of K326 is pollinated with a plant of the genetic background DH98-325-6 carrying mutations in each of the three nicotine demethylase genes. F₁individuals originating from this cross and heterozygous for each mutation are backcrossed to the recurrent parent, K326, to produce BC₁F₁progeny. BC₁F₁progeny are screened using genotyping methodologies to identify individuals heterozygous for all three mutations. A single triple heterozygous BC₁F₁plant is backcrossed to K326 to produce BC₂F₁progeny. The process of backcrossing and identification of individuals heterozygous for all three mutations is repeated through the BC₃F₁, BC₄F₁, BC₅F₁, BC₆F₁, and BC₇F₁stages. At the BC₇F₁stage, individuals heterozygous for all three mutations from each pedigree are self-pollinated to produce BC₇F₂seed. A large number of BC₇F₂progeny from each pedigree are genotyped to identify individuals homozygous for all three mutations. A single BC₇F₂plant homozygous for all three mutations is self-pollinated to produce a BC₇F₃family (K326 SRC) in which the wild-type CYP82E4, CYP82E5v2, and CYP82E10 alleles of K326 were replaced by the mutant (cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S alleles).

The male-sterile (CMS) version of K326 SRC (CMS K326 SRC) is produced by crossing a plant of CMS K326 as a female with pollen of K326 SRC to produce male-sterile plants heterozygous for all three mutations. A single male-sterile plant resulting from this cross is then subsequently backcrossed as a female to K326 SRC to produce progeny that are segregating for individuals homozygous for all three mutations. Triple homozygous individuals are identified by DNA genotyping to produce the CMS K326 SRC line. Because the line is male-sterile, it is maintained via pollination with K326 SRC.

Commercial K326 SRC is produced by pollinating plants of CMS K326 SRC with pollen of K326 SRC.

Testing of K 326 SRC

K326 SRC is evaluated for cured leaf chemistry, yield, and physical quality at three North Carolina field research locations during 2012 (Oxford, Rocky Mount, Kinston). K326 is included for comparison. The experimental design at each location is a randomized complete block design with four replications. Experimental units are single 20-plant plots. Plots are harvested and flue-cured. Plot weights are used to determine per acre yields. Cured leaf is evaluated by a former USDA tobacco grader. Fifty gram composite leaf samples are collected from each plot and analyzed for percent nicotine, nornicotine, anatabine, anabasine, and percent nicotine conversion using gas chromatography equipment.

Comparisons using the least significant difference (LSD) test indicates that K326 SRC has significantly (P<0.05) lower levels of nornicotine and percent nicotine conversion relative to K326 (Table 1). K326 and K326 SRC are not significantly different from each other for percent nicotine, yield, or cured leaf quality indices.

Analysis of NNN Levels in K 326 SRC Tobacco Filler and Cigarette Smoke

In an independent testing, K326 SRC is further evaluated for NNN levels in both tobacco filler and cigarette smoke by gas chromatography. Percent nicotine conversion in tobacco filler is also measured. K326 is included for comparison. K326 SRC has a lower level of NNN compared to K326 in both tobacco filler (˜022 versus ˜0.53 ppm) and cigarette smoke (˜0.015 versus ˜0.04 ng/mg Total Particle Matter (TPM)) (FIG. 1). Percent nicotine conversion is lower in K326 SRC tobacco filler (˜0.8%) than K326 tobacco filler (˜3.1%) (FIG. 1).

Example 2
Breeding of Homozygous cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S Mutant Plants into the K346 Flue-Cured Tobacco Cultivar

K346 SRC is a backcross-derived version of flue-cured tobacco cultivar K346 carrying introduced null mutations in three genes (CYP82E4 (SEQ ID NO: 5), CYP82E5 (SEQ ID NO: 8), and CYP82E10 (SEQ ID NO: 9)) previously documented to encode for nicotine demethylase enzymes (Lewis et al., 2010). The introduced mutations in CYP82E4 (cyp82e4 W329Stop (SEQ ID NO: 1)) and CYP82E5 (cyp82e5v2 W422Stop (SEQ ID NO: 2)) encode for premature stop codons which render the genes non-functional. The introduced mutation in CYP82E10 (cyp82e10 P381S (SEQ ID NO: 11)) does not encode for a premature stop codon, but does render the gene product non-functional for converting nicotine to nornicotine (Lewis et al., 2010). When in homozygous condition, the three mutations result in tobacco plants with (1) reduced genetic capacity to demethylate nicotine to form nornicotine, and (2) a corresponding diminished potential to accumulate N-nitrosonornicotine (NNN), a potent carcinogen found in many tobacco products. The mutation in CYP82E4 (cyp82e4 W329Stop (SEQ ID NO: 1)) also provides phenotypic stability for the “nicotine conversion” trait and eliminates the requirement to utilize the cumbersome “LC” method for reducing levels of nornicotine in tobacco cultivars.

The original tobacco cultivar K346 is a fertile inbred line. CMS K346 is a cytoplasmic male-sterile version of the original K346. To develop K346 SRC, an individual plant of K346 is initially pollinated with a plant of the genetic background DH98-325-6 carrying mutations in each of the three nicotine demethylase genes. F₁individuals originating from this cross and heterozygous for each mutation are backcrossed to the recurrent parent, K346, to produce BC₁F₁progeny. BC₁F₁progeny are screened using genotyping methodologies to identify individuals heterozygous for all three mutations. A single triple heterozygous BC₁F₁plant is backcrossed to K346 to produce BC₂F₁progeny. The process of backcrossing and identification of individuals heterozygous for all three mutations is repeated through the BC₃F₁, BC₄F₁, BC₅F₁, BC₆F₁, and BC₇F₁stages. At the BC₇F₁stage, individuals heterozygous for all three mutations from each pedigree are self-pollinated to produce BC₇F₂seed. A large number of BC₇F₂progeny from each pedigree are genotyped to identify individuals homozygous for all three mutations. A single BC₇F₂plant homozygous for all three mutations is self-pollinated to produce a BC₇F₃family (K346 SRC) in which the wild-type CYP82E4, CYP82E5v2, and CYP82E10 alleles of K346 were replaced by the mutant (cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S alleles).

The male-sterile (CMS) version of K346 SRC (CMS K346 SRC) is produced by crossing a plant of CMS K346 as a female with pollen of K346 SRC to produce male-sterile plants heterozygous for all three mutations. A single male-sterile plant resulting from this cross is then subsequently backcrossed as a female to K346 SRC to produce progeny that are segregating for individuals homozygous for all three mutations. Triple homozygous individuals are identified by DNA genotyping to produce the CMS K346 SRC line. Because the line is male-sterile, it is maintained via pollination with K346 SRC.

Commercial K346 SRC is produced by pollinating plants of CMS K346 SRC with pollen of K346 SRC.

Testing of K346 SRC

K346 SRC is evaluated for cured leaf chemistry, yield, and physical quality at three North Carolina field research locations during 2012 (Oxford, Rocky Mount, Kinston). K346 is included for comparison. The experimental design at each location is a randomized complete block design with four replications. Experimental units are single 20-plant plots. Plots are harvested and flue-cured. Plot weights are used to determine per acre yields. Cured leaf is evaluated by a former USDA tobacco grader. Fifty gram composite leaf samples are collected from each plot and analyzed for percent nicotine, nornicotine, anatabine, anabasine, and percent nicotine conversion using gas chromatography equipment.

Comparisons using the least significant difference (LSD) test indicated that K346 SRC has significantly (P<0.05) lower levels of nornicotine and percent nicotine conversion relative to K346 (Table 1). K346 and K346 SRC are not significantly different from each other for percent nicotine, yield, or cured leaf quality indices.

Example 3
Breeding of Homozygous cyp82e4 W329Stop, the cyp82e5v2 W422Stop, and cyp82e10 P381S Mutant Plants into the NC196 Flue-Cured Tobacco Cultivar

Hybrid cultivar NC196 SRC is a backcross-derived version of flue-cured tobacco cultivar NC196 carrying introduced deleterious mutations in three genes (CYP82E4 (SEQ ID NO: 5), CYP82E5 (SEQ ID NO: 8), and CYP82E10 (SEQ ID NO: 9)) previously documented to encode for nicotine demethylase enzymes (Lewis et al., 2010). The introduced mutations in CYP82E4 (cyp82e4 W329Stop (SEQ ID NO: 1)) and CYP82E5 (cyp82e5v2 W422Stop (SEQ ID NO: 2)) encode for premature stop codons which render the genes non-functional. The introduced mutation in CYP82E10 (cyp82e10 P381S (SEQ ID NO: 11)) does not encode for a premature stop codon, but does render the gene product non-functional for converting nicotine to nornicotine (Lewis et al., 2010). When in homozygous condition, the three mutations result in tobacco plants with (1) reduced genetic capacity to demethylate nicotine to form nornicotine, and (2) a corresponding diminished potential to accumulate N-nitrosonornicotine (NNN), a potent carcinogen found in many tobacco products. The mutation in CYP82E4 (cyp82e4 W329Stop (SEQ ID NO: 1)) also provides phenotypic stability for the “nicotine conversion” trait and eliminates the requirement to utilize the cumbersome “LC” method for reducing levels of nornicotine in tobacco cultivars.

The original tobacco cultivar NC196 is a hybrid generated by pollinating plants of a male-sterile breeding line CMS NCTG-61 with pollen produced by fertile breeding line NC1562-1. To develop hybrid cultivar NC196 SRC, individual plants of fertile NCTG-61 and NC1562-1 are first pollinated with a plant of the genetic background DH98-325-6 carrying mutations in each of the three nicotine demethylase genes. F₁individuals originating from each cross and heterozygous for each mutation are backcrossed to the recurrent parents (NCTG-61 or NC1562-1) to produce BC₁F₁progenies.

BC₁F₁progenies are screened using genotyping methodologies to identify individuals heterozygous for all three mutations. A single BC₁F₁plant from each pedigree is backcrossed to either NCTG-61 or NC1562-1, respectively, to produce BC₂F₁progenies. The process of backcrossing and identification of individuals heterozygous for all three mutations is repeated through the BC₃F₁, BC₄F₁, BC₅F₁, BC₆F₁, and BC₇F₁stages. At the BC₇F₁stage, individuals heterozygous for all three mutations from each pedigree are self-pollinated to produce BC₇F₂seed. A large number of BC₇F₂progeny from each pedigree are genotyped to identify individuals homozygous for all three mutations. BC₇F₂plants homozygous for all three mutations are self-pollinated to produce BC₇F₃families (NCTG-61 SRC and NC1562-1 SRC) in which the wild-type CYP82E4, CYP82E5v2, and CYP82E10 alleles of NCTG-61 and NC1562-1 are replaced by the mutant (cyp82e4 W329Stop, cyp82e5v2 W422Stop, and cyp82e10 P381S alleles).

The female parental line of NC196, CMS NCTG-61, is cytoplasmic male sterile which causes pollen to not be produced. To develop CMS NCTG-61 SRC, a plant of CMS NCTG-61 is crossed with NCTG-61 SRC to produce male-sterile plants heterozygous for all three mutations. A single male-sterile plant resulting from this cross is backcrossed as a female to NCTG-61 SRC to produce progeny that are segregating for individuals homozygous for all three mutations. Triple homozygous individuals are identified by DNA genotyping to produce the CMS NCTG-61 SRC line. Because the line is male-sterile, it is maintained via pollination with NCTG-61 SRC.

Hybrid cultivar NC196 SRC is produced by pollinating plants of CMS NCTG-61 SRC with pollen of NC1562-1 SRC.

Testing of Hybrid Cultivar NC196 SRC

Hybrid cultivar NC196 SRC is evaluated for cured leaf chemistry, yield, and physical quality at three North Carolina field research locations during 2012 (Oxford, Rocky Mount, Kinston). Hybrid cultivar NC196 is included for comparison. The experimental design at each location is a randomized complete block design with four replications. Experimental units are single 20-plant plots. Plots are harvested and flue-cured. Plot weights are used to determine per acre yields. Cured leaf is evaluated by a former USDA tobacco grader. Fifty gram composite leaf samples are collected from each plot and analyzed for percent nicotine, nornicotine, anatabine, anabasine, and percent nicotine conversion using gas chromatography equipment.

Comparisons using the least significant difference (LSD) test indicates that hybrid cultivar NC196 SRC has significantly (P<0.05) lower levels of nornicotine and percent nicotine conversion relative to NC196 (Table 1). NC196 and hybrid cultivar NC196 SRC are not significantly different from each other for percent nicotine, yield, or cured leaf quality indices,

TABLE 1

Means for Selected SRC Entries and EX12-122 entries averaged over three NC environments.

Nicotine

Nicotine
Nornicotine
Anabasine
Anatabine
Alkaloids
Sugars
Conversion

Grade
(% dry
(% dry
(% dry
(% dry
(% dry
(%
(% dry

Genotype
Yield
Cwt
Acre
Index
weight)
weight)
weight)
weight)
weight)
reducing)
weight)

NC 2326
2269
134.73
3089.22
80.6
3.581
0.064
0.027
0.203
3.876
13.59
1.76

NC 95
2519
139.44
3589.48
83.0
3.660
0.068
0.033
0.284
4.046
12.22
1.83

NCTG-61
2776
141.71
3957.14
83.8
3.659
0.069
0.032
0.258
4.019
14.16
1.85

NCTG-61
2720
142.63
3895.77
83.3
3.983
0.022
0.032
0.240
4.277
14.24
0.55

SRC

K346
2757
141.91
3948.65
84.2
3.316
0.065
0.024
0.155
3.560
14.43
1.93

CMS K346
2590
140.46
3656.09
83.2
3.425
0.019
0.027
0.187
3.659
10.85
0.56

SRC

K326
3076
143.01
4447.85
83.7
3.168
0.069
0.025
0.178
3.441
14.24
2.13

K326 SRC
3048
143.99
4436.36
84.1
3.215
0.023
0.022
0.159
3.418
15.41
0.70

NC1562-1
3052
144.81
4463.77
84.2
2.886
0.052
0.022
0.159
3.120
15.88
1.79

NC1562-1
3091
142.94
4454.60
83.8
2.950
0.016
0.022
0.170
3.158
15.73
0.55

SRC

NC 196
3042
142.76
4392.61
83.4
3.215
0.070
0.030
0.253
3.568
14.24
2.14

NC196 SRC
3155
143.23
4551.78
83.9
3.508
0.020
0.027
0.210
3.765
15.56
0.57

LSD 0.05
246
5.47
401.23
2.7
0.343
0.010
0.003
0.027
0.367
2.22
0.37

Deposit Information

A deposit of the proprietary inbred plant lines and hybrid disclosed above and recited in the appended claims have been made with American Type Culture Collection (ATCC), 10801 University Boulevard, Manassas, Va. 20110. The date of deposit for K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NC196 SRC, and CMS NCTG-61 SRC was Jan. 17, 2013. The date of deposit for NCTG-61 SRC was May 7, 2013. The deposits of 2500 seeds for each variety was taken from the same deposits maintained since prior to the filing date of this application. Upon issuance of a patent, all restrictions upon the deposits will be removed, and the deposits are intended to meet all of the requirements of 37 C.F.R. §1.801-1.809. The ATCC has issued the following accession numbers: ATCC Accession No. PTA-13455 for K326 SRC; ATCC Accession No. PTA-13454 for CMS K326 SRC; ATCC Accession No. PTA-13453 for K346 SRC; ATCC Accession No. PTA-13452 for CMS K346 SRC; ATCC Accession No. PTA-13457 for NC1562-1 SRC; ATCC Accession No. PTA-13456 for CMS NCTG-61 SRC; ATCC Accession No. PTA-120314 for NCTG-61 SRC; and ATCC Accession No. PTA-13458 for hybrid NC196 SRC. These deposits will be maintained in the depository for a period of 30 years, or 5 years after the last request, or for the effective life of the patent, whichever is longer, and will be replaced as necessary during that period.

	Number	Date	Country
	61830908	Jun 2013	US
	61751720	Jan 2013	US

TOBACCO INBRED PLANTS K326 SRC, CMS K326 SRC, K346 SRC, CMS K346 SRC, NC1562-1 SRC, NCTG-61 SRC, CMS NCTG-61 SRC AND HYBRID NC196 SRC

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