EP0395398A2
Expired / Lapsed
EP
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Amplification of nucleic acid sequences using random sequence oligonucleotides as primers
Assignee: Life Technologies Inc
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Filing: 1990-04-26
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Granted: —
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Expires: 2010-04-26
Abstract
Abstract A process for substantially amplifying template nucleic acid present in a sample is described, wherein the amplification may be performed without the need for prior knowledge of specific sequences. The process comprises apposition of random oligonucleotide primers to the template nucleic acid under conditions such that extension products of the primers are synthesised which are complementary to the template nucleic acid.
Inventors
- 👤 James L. Hartley
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-08 20:27
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/70; C12Q1/701; C12Q1/708; C12Q1/68; C12Q1/6844; C12Q1/6858; Y; Y10; Y10S; Y10S435/00; Y10S435/81; Y10T; Y10T436/00; Y10T436/14; Y10T436/142222; Y10T436/143333
Claims Analysis
Total Claims: 24
Claim 1
Independent Claim
28 words
A process for substantially amplifying a template nucleic acid sequence in a sample, the process comprising amplifying the nucleic acid in a randomly primed and template dependent manner.
Claim 2
Dependent Claim
28 words
A process for substantially amplifying a template nucleic acid sequence in a sample, the process comprising amplifying the nucleic acid in a randomly primed and template dependent manner.
Claim 3
Dependent Claim
69 words
A process for amplifying a nucleic acid sequence, such as in a sample, the process comprising: (a) hybridising a random oligonucleotide primer to a template nucleic acid sequence; (b) synthesising a substantially complementary nucleic acid sequence from the primer; (c) separating the nucleic acid and substantially complementary nucleic acid sequences; (d) hybridising a random oligonucleotide primer to the nucleic acis or complementary nucleic acid sequence; and (e) repeating (b).
Claim 4
Dependent Claim
69 words
A process for amplifying a nucleic acid sequence, such as in a sample, the process comprising: (a) hybridising a random oligonucleotide primer to a template nucleic acid sequence; (b) synthesising a substantially complementary nucleic acid sequence from the primer; (c) separating the nucleic acid and substantially complementary nucleic acid sequences; (d) hybridising a random oligonucleotide primer to the nucleic acis or complementary nucleic acid sequence; and (e) repeating (b).
Claim 5
Dependent Claim
46 words
A process for detecting a papilloma virus in a sample the process comprising: (a) substantially amplifying a papilloma virus nucleic acid sequence in a sample wherein the substantial amplification comprises a randomly primed template dependent synthesis of papilloma virus DNA; and (b) detecting the papilloma virus.
Claim 6
Dependent Claim
46 words
A process for detecting a papilloma virus in a sample the process comprising: (a) substantially amplifying a papilloma virus nucleic acid sequence in a sample wherein the substantial amplification comprises a randomly primed template dependent synthesis of papilloma virus DNA; and (b) detecting the papilloma virus.
Claim 7
Dependent Claim
63 words
A process for substantially amplifying a template nucleic acid sequence in a sample in a randomly primed and template dependent manner the process comprising: (a) priming a template nucleic acid sequence with a random oligonucleotide primer; and (b) incubating the template nucleic acid sequence and the random oligonucleotide primer in the presence of an inducing agent, a strand displacement agent and triphosphate substrates.
Claim 8
Dependent Claim
63 words
A process for substantially amplifying a template nucleic acid sequence in a sample in a randomly primed and template dependent manner the process comprising: (a) priming a template nucleic acid sequence with a random oligonucleotide primer; and (b) incubating the template nucleic acid sequence and the random oligonucleotide primer in the presence of an inducing agent, a strand displacement agent and triphosphate substrates.
Claim 9
Dependent Claim
24 words
A process as claimed in any of claims 1 to 4 wherein randomly primed amplification occurs with a primer that is 4-20 bases long.
Claim 10
Dependent Claim
24 words
A process as claimed in any of claims 1 to 5 wherein randomly primed amplification occurs with a primer that is 8 bases long.
Claim 11
Dependent Claim
39 words
A process as claimed in any of claims 4 to 6 wherein the inducing agent is E. coli DNA polymerase I, the Klenow fragment of E. coli DNA polymerase I, bacteriophage T5 DNA polymerase, or bacteriophage T7 DNA polymerase.
Claim 12
Dependent Claim
30 words
A process as claimed in any of claims 1 to 7 wherein the sample is derived from a biological source such as a human, animal, plant, bacterial or viral source.
Claim 13
Dependent Claim
19 words
A process as claimed in claim 8 wherein the viral source is human papilloma virus DNA or HIV DNA.
Claim 14
Dependent Claim
45 words
A process as claimed in any of claims 1 to 9 wherein the sample is derived from a tissue, fluid or secretion of the biological source, such as blood, stool, sera, urine, saliva, tears, biopsy tissue sample, histology tissue sample, PAP smear, mole or wart.
Claim 15
Dependent Claim
26 words
A process as claimed in any of claims 4 to 10 wherein the inducing agent and the strand displacement agent are activities of the same protein.
Claim 16
Dependent Claim
73 words
A process for substantially amplifying a template nucleic acid sequence in a sample in a randomly primed and template dependent manner, the process comprising: (a) priming a template nucleic acid sequence with one or more random oligonucleotide primers wherein the primer(s) consist of 6-mers to 10-mers; and (b) incubating the template nucleic acid strand and the random oligonucleotide primer(s) in the presence of the Klenow fragment of DNA Polymerase I and triphosphate substrates.
Claim 17
Dependent Claim
73 words
A process for substantially amplifying a template nucleic acid sequence in a sample in a randomly primed and template dependent manner, the process comprising: (a) priming a template nucleic acid sequence with one or more random oligonucleotide primers wherein the primer(s) consist of 6-mers to 10-mers; and (b) incubating the template nucleic acid strand and the random oligonucleotide primer(s) in the presence of the Klenow fragment of DNA Polymerase I and triphosphate substrates.
Claim 18
Dependent Claim
41 words
A process for detecting the presence and/or amount of a substance which comprises a nucleic acid sequence, the process comprising amplifying a template nucleic acid sequence as claimed in any of claims 1 to 12, and detecting the amplified nucleic acid.
Claim 19
Dependent Claim
13 words
A process as claimed in claim 13 wherein the substance is papilloma virus.
Claim 20
Dependent Claim
53 words
A kit for amplifying a nucleic acid sequence, the kit comprising a carrier having: (a) a container containing a random oligonucleotide primer; (b) a container containing an inducing agent; (c) a container containing triphosphate substrates; (d) a container containing a buffer for reconstituting or diluting any or all of components (a) to (c).
Claim 21
Dependent Claim
53 words
A kit for amplifying a nucleic acid sequence, the kit comprising a carrier having: (a) a container containing a random oligonucleotide primer; (b) a container containing an inducing agent; (c) a container containing triphosphate substrates; (d) a container containing a buffer for reconstituting or diluting any or all of components (a) to (c).
Claim 22
Dependent Claim
13 words
A kit as claimed in claim 15 further comprising a strand displacement agent.
Claim 23
Dependent Claim
24 words
A kit as claimed in claim 15 or 16 further comprising a container containing a probe capable of identifying a bacterial or viral species.
Claim 24
Dependent Claim
23 words
A kit as claimed in claim 17 wherein the probe is a human papilloma virus probe, a HIV probe, or a Salmonella probe.
References Cited (Backward Citations)
No citation records indexed.
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