Patent Grant
PP20149
The invention provides a new distinct somaclone of rose scented geranium P. graveolens christened as ‘Parimal’ characterized by distinct morphology and improved oil yield determining parameters. The said novel somaclone has been developed from callus through in vitro tissue culture technique, without enforced mutagenesis. The plant possesses characteristic leaf morphology, vigorous growth, improved oil content per plant, herb yield and oil yield. The oil produced by this somaclone resembles the oil produced by the commercial type of bourbon geranium oil in that has equivalent ratio of citronellol and geraniol.
The present invention relates to a new and distinct somaclone of rose scented geranium Pelargonium graveolens a member of the family Geraniaceae, which is a stable somaclone of the Indian cultivar Hemanti. The oil of geranium finds extensive use in perfumery, cosmetics, food and pharmaceutical industries (Narayana, 1986). Geranium was introduced in India in the beginning of this century and since then its cultivation and production of oil remained restricted to the high altitude regions of Ootacamund, Kodaikanal and Yercaud areas in South India (Narayana, 1986). India produces hardly 5 tonnes of geranium oil annually as against its own requirement of approximate 100 tonnes per year (Narayana 1986). The production of oil has declined gradually over the years and much of the requirement is met by imports. This may be due to narrow genetic base, resulting partially from sexual infertility, vegetative mode of propagation and restricted area of cultivation.
In India, two cultivars of geranium are mainly known; these are the types that produce Algerian or Tunisian quality and the Bourbon or the Reunion quality of geranium oils; these are given the names Pelargonium×graveolens cv Hemanti and Pelargonium×graveolens cv Bipuli, respectively. The cultivar Hemanti was found to be more adaptive than the cultivar Bipuli both in the hilly regions as well as in the unconventional areas of the plains. On the other hand, the cultivar Bipuli scored better than the Hemanti mainly because of its better oil yield and improved organoleptic value of the oil. The perfumery value of geranium oil is mainly determined by the ratio of citronellol and geraniol and almost equal contents of these two major terpenoid components is generally considered to be the characteristic of the best quality geranium oil.
In an attempt to extend the area of cultivation of geranium to the Indo-gangetic plains, the better adaptive Hemanti cultivar was introduced in the Lucknow conditions in India. Subsequently, efforts were made through different biotechnological methods to develop clones with high content and desirable composition of the oil. The calliclone ‘Parimal ’evolved through this invention represents improvement over the wild type Hemanti parental cultivar both in terms of quality and quantity determining traits of the essential oil. This particular somaclone, being consistently conditions, can offer an improved cultivar for commercial exploitation.
The main objective of the inventin is to develop useful somaclonal variants of rose scented geranium christened as ‘Parimal ’which are distinct, stable and uniform through successive generations.
Another object is to develop somaclonal variants yielding high quality and quantity of essential oil.
Accordingly the present invention provides a novel variety of rose scented geranium called ‘Parimal ’said plant developed through in vitro tissue culture methods from callus cultures. The invention further provides a useful somaclonal variant ‘Parimal’of rose scented geranium (Pelargonium graveolens cv. Hemanti) yielding high quality and quantity of essential oil.
In accordance with the above and other objects, the invention provides a novel somaclone of rose scented geranium called ‘Parimal’. The said plant has been developed though in vitro tissue cultured on modified Murashige and Skoog's medium (MS), 1962.
The invention further provides a useful somaclonal variant ‘Parimal’ of rose scented geranium (Pelargonium graveolens cv. Hemanti) having the following morphological characteristics.
This invention further provides the useful somaclonal variant “Parimal” of rose scented geranium (P. graveolens cv. Hemanti) having the following essential oil quality determining characteristics. The average and mean percentage of the respective constituents of the essential oil are provided hereinbelow:
This invention further provides the said somaclone “Parimal” which exhibited uniqueness with respect to the following RAPD primers:MAP-05, MAP-12, MAP-15 and MAP-16.
The methods and examples for the development of the said novel plant “Parimal” are provided herein below, for illustration of the invention and should not be construed as limitations on the concept of the invention. Modifications that may be apparent to those skilled in the art are deemed to fall within the scope of the present invention.
The invention is further illustrated with reference to the following drawings wherein:
FIG. (a-d): Variation in the RAPD profiles of wild type parent (C) with that of the selected somaclone “Parimal” (marked with (→)arrow); a:primer MAP-5; b:primer MAP-12; c:primer MAP-15; d: primer MAP-16.
Methods for development of the novel somaclonal variant “Parimal”:
Healthy robust shoots, raised by subculturing the organogenic calli on modified MS medium with concentrations of BAP and NAA, were transferred to half strength MS medium with a concentration of indole butyric acid (UBA) for rhizogenesis. The stem explants were cultured on modified Murashige and Skoog's medium (MS), 1962 containing the following (in mg/L)-NH4NO3 (1650), KNO3 (1900), CaCl2.2H2O (400), MgSO4.7H2O (370), KH2PO4 (170), Na2EDTA.2H2O (7.2), FeSO4.7H2O (27.8), MnSO4.4H2O (22.3), ZnSO4.7H2O (8.6), H3BO3 (6.2), KI (0.83), Na2MoO4.2H2O (0.25), CuSO4.5H2O (0.025), CoCl2.6H2O (0.025), Glycine (2.0), Nicotinic acid (0.5), Pyrodoxine HCl (0.5) and Thiamine HCl (0.1) with proportion of naphthelene acetic acid (NAA) and Kinetin (Kn)[2:1]. The best organogenesis (85%) could be achieved by substituting Kinetin with 6-Benzylaminopurine (BAP).
The rooted plants were initially hardened under glass house conditions and were subsequently transferred to the field in the month of October 1996, in replicated trials. Based on the superior performance for vigorous growth resulting from extensive branching and enlarged canopy size, comparatively better oil yield over that of the control wild type cultivar and distinctively improved oil quality in terms of the major terpenoid constituents, the plant of this invention (Somaclone “Parimal”) was selected for further observation and evaluations. During the monsoon seasons of 1996-97, this selected somaclone along with control parent were maintained under glass-house conditions through stem-cuttings which also maintained clonal purity. The selected clone “Parimal” was grown in field in replicated trials along with its parent Hemanti cultivar for two consecutive winter-summer seasons of 1997-98 and 1998-99, while during the intermediate monsoon season, they were maintained under glass-house conditions. The clone “Parimal” maintained its vigorous growth through extensive branching, better oil content and improved oil quality over the three consecutive years. The characteristics of this particular clone (in terms essential oil quantity and quality determining traits) in relation to that of the control parent and other clones have been represented in Tables-2 and 3. The results clearly demonstrate the outstanding performance of clone “Parimal” over that of the control wild type parent and other selected clones under Lucknow field conditions, indicating the feasibility of its commercial cultivation in the indo-gangetic plains. It is pertinent to mention ere that the oil profile of somaclone “Parimal” is superior than that of the wild type parent in having equal content of citronellol and geraniol which is a critical characteristic of best quality commercial geranium oil. The field grown plant of “Parimal” having large round canopy is shown in FIG. 1 and
Evidences of uniformity and stability
The somaclone “Parimal” has remained stable and uniform for its morphological characters and showed consistency in performance for various oil yield and quality determining attributes during its field evaluation under Lucknow conditions over three consecutive years of 1996-97, 1997-98 and 1998-99 are depicted in (Tables 1-3).
Morphological description of the somaclone “Parimal” and its wild type parent
Type and frequency of variants during multiplication and maintenance of the somaclone
No variants has been recovered during multiplication, maintenance and testing of the somaclone “Parimal” in experimental plots, screened carefully in 1996-97, 1997-98 and 1998-99 winter-summer cropping seasons. The three year's observations on somaclone “Parimal” were statistically analysed for significance of differences in the morphological oil yield determining attributes and the concentrations of 9 geranium oil quality determining terpenoid constituents. The intraclonal variation in clone “Parimal” was found statistically insignificant over three year's field trial.
Statement of distinction
The somaclones “Parimal” are distinct in having large round canopy, less dentated round leaf morphology, extensive branching and absence of flowering (Table 1). Its oil profile is distinct from the wild type parent cultivar in having higher proportion of geraniol and lower proportion of citronellol, lower content of cis+trans rose oxide and citronellyl formate, 6,9-guadiene, higher content of linalool, geranyl formate and 10-epi-y endesmol (Table-2). Its RAPD profile was different from the wild type parent in certain aspects while had certain features common in both.
Characterization through RAPD markers
Using 13 random primers (MAP 01 to MAP 16) with following sequence description a comparison was made for somaclone “Parimal” and its wild type parent to establish the relatedness between them.
This analysis could narrow down with usage of six MAP primers (MAP 04, MAP 05, MAP 10, MAP 14, MAP 15, MAP 16) which in turn were utilized for genotypic pair analysis in order to assess genetic/molecular distances. The following table 4 gives the similarity index between the two genotypes in paired test.
It is evident from similarity indices (paired test) that somaclone “Parimal” and wild type parent are genetically distinguishable (Table 4). Further, the primer MAP 10, MAP 14 showed least similarity while primers MAP 04 and MAP15 showed close relatedness. The primers MAP05 and MAP16 showed intermediate degree of similarity. These observations on percent similarity confirm the relatedness of somaclone “Parimal” with the wild type parent, at the same time highlight the distinctness of the former at the genetic level. The complete RAPD profile for PCR amplified DNA segments using 12 primers is available providing evidence of distinctiveness of the clone with respect to the control parent as shown in FIG. 4.
Advantages:
| Number | Name | Date | Kind |
|---|---|---|---|
| PP17422 | Westhoff | Feb 2007 | P2 |
