Patent EP0369775A2 successfully refreshed from Google Patents.
EP0369775A2
Expired / Lapsed
EP
⏱️ 0.0 years remaining
Compositions and process for amplifying and detecting nucleic acid sequences
Assignee: Lifecodes Corp
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Filing: 1989-11-15
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Granted: —
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Expires: 2009-11-15
Abstract
Abstract Compositions and methods useful for detecting, amplifying and/or characterizing polynucleotides of interest are disclosed herein. Specifically, this disclosure teaches how to make and use a mobile promoter structure comprising a transcription promoter and a probe sequence. The probe sequence enables the mobile promoter structure to select via hybridization a specific polynucleotide of interest. The promoter enables the transcription of the target polynucleotide. The method of mobile promoter structure-directed transcription can be applied to detect, and/or characterize pathogens, disease states, and genetic traits. While a single mobile promoter structure-directed transcription enables linear amplification, the process can be used in combination with other processes to bring about exponential amplification.
Inventors
- 👤 Zvi G. Loewy
- 👤 Howard J. Baum
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-30 14:13
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6876; C12Q1/6844; C12Q1/6865; C12Q2600/00; C12Q2600/158
Claims Analysis
Total Claims: 18
Claim 1
Independent Claim
86 words
A mobile promoter structure comprising: (a) a first polynucleotide sequence comprising, sequentially-linked, a plus promoter sequence, an optional plus bridge sequence, and a probe sequence at the 3′-terminus of the first polynucleotide sequence; and (b) a second polynucleotide sequence comprising a minus promoter sequence complementary to the plus promoter sequence, and an optional minus bridge sequence complementary to the optional plus bridge sequence, wherein the 5′-terminus of the second polynucleotide sequence is the 5′-terminus of the minus promoter sequence or, if present, the minus bridge sequence.
Claim 2
Dependent Claim
86 words
A mobile promoter structure comprising: (a) a first polynucleotide sequence comprising, sequentially-linked, a plus promoter sequence, an optional plus bridge sequence, and a probe sequence at the 3′-terminus of the first polynucleotide sequence; and (b) a second polynucleotide sequence comprising a minus promoter sequence complementary to the plus promoter sequence, and an optional minus bridge sequence complementary to the optional plus bridge sequence, wherein the 5′-terminus of the second polynucleotide sequence is the 5′-terminus of the minus promoter sequence or, if present, the minus bridge sequence.
Claim 3
Dependent Claim
22 words
A mobile promoter structure according to claim 1, wherein the plus bridge sequence is present and is at least one nucleotide long.
Claim 4
Dependent Claim
19 words
A mobile promoter structure according to claim 2, wherein the plus bridge sequence is at least 6 nucleotides long.
Claim 5
Dependent Claim
13 words
A mobile promoter structure according to claim 1, which is 5′-AATTAACCCTCACTAAAGGGAACCTGCGCAAGGAACGCCCGTCGTGGC-3′ 3′-TTAATTGGGAGTGATTTCCCTTG -5′.
Claim 6
Dependent Claim
29 words
An annealed complex comprising a mobile promoter structure according to any of claims 1 to 4, and a target polynucleotide of which at least the 3′-terminal portion is single-stranded.
Claim 7
Dependent Claim
28 words
An annealed complex according to claim 5, wherein the 5′-terminus of the second polynucleotide sequence of the mobile promoter and the 3′-terminus of the target polynucleotide are ligated.
Claim 8
Independent Claim
55 words
A method of transcribing a target polynucleotide sequence, comprising: (a) incubating a mobile promoter structure according to any of claims 1 to 4 with the target polynucleotide, under conditions such that they form an annealed complex; and (b) incubating the annealed complex under conditions which give transcription under the direction of the mobile promoter structure.
Claim 9
Dependent Claim
30 words
A method according to claim 7, wherein the target polynucleotide is part of a double-stranded polynucleotide, and which comprises the preliminary additional step of separating the two strands by denaturation.
Claim 10
Dependent Claim
33 words
A method according to claim 7 or claim 8, wherein step (b) is conducted in the presence of an RNA polymerase selected from T3 RNA polymerase, T7 RNA polymerase and SP6 RNA polymerase.
Claim 11
Independent Claim
36 words
A method according to any of claims 7 to 9, which additionally comprises, after step (a) but prior to step (b), ligating the 5′-terminus of the second polynucleotide sequence and the 3′-terminus of the target polynucleotide.
Claim 12
Dependent Claim
64 words
A method of transcribing polynucleotide sequences, comprising performing n (n being an integer of at least two) cycles of mobile promoter structure-directed transcription, wherein each cycle comprises a method according to claim 10, and wherein, in the n th cycle, the probe sequence anneals with the 3′ region of a target polynucleotide which is complementary to the target polynucleotide of the (n-1) th cycle.
Claim 13
Independent Claim
37 words
A method of transcribing and amplifying a target polynucleotide sequence, comprising performing alternating cycles of mobile promoter structure-directed transcription according to the method of any of claims 7 to 10, and reverse transcription by primer extension synthesis.
Claim 14
Dependent Claim
17 words
A method according to claim 12, which comprises at least 2 cycles of mobile promoter structure-directed transcription.
Claim 15
Independent Claim
18 words
A method according to any of claims 11 to 13, wherein all of the plus promoters are identical.
Claim 16
Independent Claim
27 words
A method according to any of claims 7 to 14, which additionally comprises determining the size of, and/or sequencing, a transcription product, thereby characterising the target polynucleotide.
Claim 17
Independent Claim
58 words
A method for transcribing an RNA template which comprises (1) annealing an RNA sequence with a mobile promoter, e.g. according to any of claims 1 to 4; (2) ligating the resulting complex, e.g. with T4 RNA polymerase; and (3) incubating the ligated annealed complex with an RNA polymerase, e.g. T3 RNA polymerase, in the presence of ribonucleoside triphosphates.
Claim 18
Dependent Claim
23 words
A method according to claim 16, wherein the RNA sequence is complementary to the 270 base sequence of the neo gene of Tn5.
References Cited (Backward Citations)
No citation records indexed.
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