Claims
- 1. A purified, recombinantly produced enzyme preparation derived from Thermus thermophilus which has the following characteristics:
- (1) it catalyzes the combination of nucleoside triphosphates to form a nucleic acid strand complementary to a template nucleotide sequence;
- (2) it is thermostable;
- (3) it has a temperature optimum for polymerization activity between 50.degree. and 90.degree. C.;
- (4) it is able to function effectively in a polymerase chain reaction, wherein said reaction includes repeated exposure to a denaturation temperature of 90.degree.-100.degree. C.;
- (5) it is free of nucleic acid encoding other Thermus thermophilus proteins;
- (6) it is free of contaminating Thermus thermophilus proteins, and;
- (7) wherein said enzyme contains an amino acid sequence selected from the group consisting of Seq ID No. 31, the sequence that is amino acids 10 to 834 of Seq ID No. 31, the sequence that is amino acids number 80 to 834 of Seq ID No. 31, and the sequence that is approximately amino acids number 279 to 834 of Seq ID No. 31.
- 2. The purified enzyme of claim 1 wherein said enzyme consists of amino acids 80-834 of Seq ID No. 31.
- 3. The purified enzyme of claim 1 wherein said enzyme consists of amino acids 10-834 of Seq ID No. 31.
- 4. The purified enzyme of claim 1 wherein said enzyme consists of amino acids 279-834 of Seq ID No. 31.
- 5. A recombinantly produced enzyme preparation derived from Thermus thermophilus which comprises a DNA polymerase and has the following characteristics:
- (1) it catalyzes the combination of nucleoside triphosphates to form a nucleic acid strand complementary to a template nucleotide sequence;
- (2) it is thermostable;
- (3) it has a temperature optimum for polymerization activity between 50.degree. and 90.degree. C.;
- (4) it is able to function effectively in a polymerase chain reaction, wherein said reaction includes repeated exposure to a denaturation temperature of 90.degree.-100.degree. C.;
- (5) it has reverse transcriptase activity;
- (6) it is free of nucleic acids encoding other Thermus thermophilus proteins; and
- (7) it is free of contaminating Thermus thermophilus proteins, wherein said DNA polymerase consists of an amino acid sequence that is SEQ ID NO: 31.
CROSS-REFERENCE
This application is a divisional, of application Ser. No. 08/384,490 filed Feb. 6, 1995, now U.S. Pat. No. 5,618,711, which is a continuation of Ser. No. 08/148,133, filed Nov. 2, 1993 now abandoned, which is a continuation of U.S. Ser. No. 07/880,478, filed May 6, 1992, now abandoned, which is a continuation of U.S. Ser. No. 07/455,967, filed Dec. 22, 1989, now abandoned, which is a continuation-in-part of U.S. Ser. No. 07/143,441, filed Jan. 12, 1988, now abandoned, which is a continuation-in-part of U.S. Ser. No. 07/063,509, filed Jun. 17, 1987, which issued as U.S. Pat. No. 4,889,818, which is a continuation-in-part of U.S. Ser. No. 06/899,241, filed Aug. 22, 1986, now abandoned.
US Referenced Citations (5)
Foreign Referenced Citations (3)
Number |
Date |
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0258017 |
Mar 1988 |
EPX |
0359006 |
Aug 1989 |
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8906691 |
Jul 1989 |
WOX |
Non-Patent Literature Citations (14)
Entry |
Sakaguchi and yajima, May, 1974, "Thermophilic and Stable DNA Polymerase From Thermus Thermophilus" Fed. Proceeds. 33(5):1492 (Abstract No. 1519). |
Johnson et al., November, 1985, "Isolation of the Gene Encoding Yeast DNA Polymerase I" Cell 43:369-377. |
Ruttimann et al., 1985, "DNA Polymerases From the Extremely Thermophilic Bacterium Thermus Thermophilus HB-8" Eur. J. Biochem. 149:41-46. |
Bechtereva et al., 1989, "DNA Sequencing With Thermostable Tet DNA Polymerase From Thermus Thermophilus" Nucleic Acids Research 17(24):10507. |
Lawyer et al., Apr., 1989, "Isolation, Characterizations, and Expression in Escherichia coli of the DNA Polymerase Gene from Thermus Aquaticus" J. Biological Chemistry 264(11):6427-6437. |
Sambrook et al., Molecular Cloning: Laboratory Manual, second edition, vol. 2, Cold Spring Harbor Laboratory Press, N.Y. (1989) pp. 13.3-13.10 "Sequencing Techniques and Strategies". |
Chien et al., Sep., 1976, "Deoxyribonucleic Acid Polymerase From the Extreme Thermophilie Thermus aquaticus" J. Bacteriology 127(3):1550-1557. |
Stoesz and Lumry, 1978, "Refolding Transition of alpha-Chymotrypsin: pH and Salt Dependence" Biochemistry 17(18):3693-3699. |
Hon-nami and Oshima, 1979, "Denaturation of Thermophilic Ferricytochrome c-552 by Acid, Guanidine Hydrochloride, and Heat" Biochemistry 18(25):5693-5697. |
Kaledin et al., Apr., 1980, "Isolation and Properties of DNA Polymerase From Extremely Thermophilic Bacterium Thermus aquaticus YT1" Biokhimiya 45(4):644-651. |
Elie et al., 1989, "Thermostable DNA Polymerase From the Archaebacterium Sulfolobus Acidocaldrius Purification, Characterization and Immunological Properties" Eur. J. Biochem. 178:619-626. |
Finnzymes Oy Tth DNA Polymerase Produce Sheets (no date available). |
Toyobo Co., Ltd. Tth DNA Polymerase Product Sheets, 1988. |
USB Tth DNA Polymerase Product Sheet, 1989. |
Divisions (1)
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384490 |
Feb 1995 |
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Continuations (3)
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148133 |
Nov 1993 |
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880478 |
May 1992 |
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455967 |
Dec 1989 |
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Continuation in Parts (3)
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143441 |
Jan 1988 |
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63509 |
Jun 1987 |
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899241 |
Aug 1986 |
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