US4921794A
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T7 DNA polymerase
Assignee: Harvard University
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Filing: 1988-12-05
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Granted: 1990-05-01
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Expires: 2008-12-05
Abstract
Abstract This invention relates to T7-type DNA polymerases and methods for amplification of DNA, for example by polymerase chain reaction.
Inventors
- 👤 Stanley Tabor
- 👤 Charles C. Richardson
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-04 21:13
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6844; C12Q1/686; C12N; C12N9/00; C12N9/10; C12N9/12; C12N9/1241; C12N9/1252; C12P; C12P19/00; C12P19/26; C12P19/28; C12P19/30; C12P19/34
Claims Analysis
Total Claims: 27
Claim 1
Independent Claim
85 words
A method of amplification of a DNA sequence comprising annealing a first and second primer to opposite strands of a double stranded DNA sequence and incubating the annealed mixture with a T7- type DNA polymerase having less than 500 units of exonuclease activity per mg of polymerase, wherein said first and second primers anneal to opposite strands of said DNA sequence with their 3' ends directed towards each other after annealing, and with the DNA sequence to be amplified located between the two annealed primers.
Claim 2
Independent Claim
85 words
A method of amplification of a DNA sequence comprising annealing a first and second primer to opposite strands of a double stranded DNA sequence and incubating the annealed mixture with a T7- type DNA polymerase having less than 500 units of exonuclease activity per mg of polymerase, wherein said first and second primers anneal to opposite strands of said DNA sequence with their 3' ends directed towards each other after annealing, and with the DNA sequence to be amplified located between the two annealed primers.
Claim 3
Dependent Claim
36 words
The method of claim 1 wherein said method further comprises, after said incubating step, denaturing the resulting DNA, annealing said first and second primers to said resulting DNA and incubating the annealed mixture with said polymerase.
Claim 4
Dependent Claim
22 words
The method of claim 2 wherein said steps of denaturing, then annealing and then incubating are repeated from 10 to 40 times.
Claim 5
Dependent Claim
18 words
The method of claim 1 wherein said exonuclease activity is less than 1 unit per mg of polymerase.
Claim 6
Dependent Claim
33 words
The method of claim 1, said T7- type DNA polymerase being selected from the group consisting of the DNA polymerases of phage T7, T3, Φ, ΦII, H, W31, gh-1, Y, A1122 and Sp6.
Claim 7
Dependent Claim
14 words
The method of claim 1 wherein said polymerase is non-discriminating for dideoxy nucleotide analogs.
Claim 8
Dependent Claim
24 words
The method of claim 1 wherein said polymerase is a modified polymerase having less than 50 units of exonuclease activity per mg of polymerase.
Claim 9
Dependent Claim
20 words
The method of claim 7 wherein said modified polymerase has less than 1 unit of activity per mg of polymerase.
Claim 10
Dependent Claim
20 words
The method of claim 7 wherein said modified polymerase has less than 0.1 unit of activity per mg of polymerase.
Claim 11
Dependent Claim
13 words
The method of claim 1 wherein said polymerase has no detectable exonuclease activity.
Claim 12
Dependent Claim
16 words
The method of claim 1 wherein said first and second primers include 10 bases or more.
Claim 13
Dependent Claim
16 words
The method of claim 1 wherein said first and second primers include 4 bases or more.
Claim 14
Dependent Claim
16 words
The method of claim 1 wherein said first and second primers comprise 4 to 20 bases.
Claim 15
Dependent Claim
27 words
The method of claim 1 wherein said polymerase is non-discriminating for nucleotide analogs and said first and second primers are single-stranded RNA or DNA comprising 4-10 bases.
Claim 16
Dependent Claim
17 words
The method of claim 1 wherein said first and second primers are single stranded DNA or RNA.
Claim 17
Independent Claim
54 words
A method of amplification of a DNA sequence comprising annealing a first and second primer to opposite strands of a double stranded DNA sequence and incubating the annealed mixture with a processive T7-type DNA polymerase, having less than 50% of the exonuclease activity of the naturally occurring level of exonuclease activity of said polymerase.
Claim 18
Independent Claim
54 words
A method of amplification of a DNA sequence comprising annealing a first and second primer to opposite strands of a double stranded DNA sequence and incubating the annealed mixture with a processive T7-type DNA polymerase, having less than 50% of the exonuclease activity of the naturally occurring level of exonuclease activity of said polymerase.
Claim 19
Dependent Claim
27 words
The method of claim 16 wherein said polymerase has less than 1% of the exonuclease activity of the naturally occurring level of exonuclease activity of said polymerase.
Claim 20
Dependent Claim
15 words
The method of claim 16 wherein said T7- type DNA polymerase is T7 DNA polymerase.
Claim 21
Dependent Claim
29 words
The method of claim 18 wherein said T7 DNA polymerase has less than 1% of the exonuclease activity of the naturally occurring level of exonuclease activity of said polymerase.
Claim 22
Independent Claim
64 words
A method of amplification of a DNA sequence comprising annealing a first and second primer to opposite strands of a double stranded DNA sequence and incubating the annealed mixture with a processive T7- type DNA polymerase, said polymerase having a level of exonuclease activity which is sufficiently low to permit the nucleotide base sequence of the DNA molecule to be determined with said polymerase.
Claim 23
Independent Claim
64 words
A method of amplification of a DNA sequence comprising annealing a first and second primer to opposite strands of a double stranded DNA sequence and incubating the annealed mixture with a processive T7- type DNA polymerase, said polymerase having a level of exonuclease activity which is sufficiently low to permit the nucleotide base sequence of the DNA molecule to be determined with said polymerase.
Claim 24
Dependent Claim
15 words
The method of claim 20 wherein said T7- type DNA polymerase is T7 DNA polymerase.
Claim 25
Dependent Claim
30 words
The method of claim 20 wherein said processive DNA polymerase has less than one percent of the exonuclease activity of the naturally occurring level of exonuclease activity of said polymerase.
Claim 26
Dependent Claim
13 words
The method of claim 22 wherein said polymerase has no detectable exonuclease activity.
Claim 27
Dependent Claim
14 words
The method of claim 22 or 23 wherein said polymerase is T7 DNA polymerase.
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