Biomerieux, Inc.

Pierre Levasseur

Philippe L Cleuziat

Francois Mallet

Francoise Guillou-Bonnici

William McAllister

Francedillaois Mallet

Francedillaoise Guillou-Bonnici

The extension of primers capable of hybridizing to the nucleic acid of interest results in a process for generating single-stranded polynucleotides. The process includes contacting the nucleic acid of interest under conditions allowing extension of the primers with a first primer and a second primer capable of hybridizing to a first region and a second region, respectively, of the nucleic acid, wherein the second region overlaps with the first region.

Method for nucleic acid amplification by transcription using displacement, and reagents and kit therefor

An oligonucleotide is intended to be used as a promoter non-template strand in the transcription of a sequence of a nucleotide target in the presence of a phage RNA polymerase. The phage RNA polymerase has specific natural promoters containing a consensus sequence from at least position -17 to position -1. The oligonucleotide contains a core sequence flanked at at least one of its ends by a nucleotide sequence capable of hybridization with a sequence of the target. The core sequence contains a sequence of 6 to 9 consecutive nucleotides from the region -12 to -4 of the non-template strand of the specific promoter, or a sufficiently homologous sequence to enable the functionality of the RNA polymerase to be retained. One flanking sequence is complementary to a first region of the target, and a second flanking sequence, when present, is complementary to a second region of the target, the first and second regions being separated on the target by a sequence having a number of nucleotides equal to the number of nucleotides in the core sequence. The number of nucleotides in the flanking region, or the sum of the number of nucleotides in the flanking regions, is at least sufficiently high for the nucleotide to be able to hybridize with the target at the temperature of use of the RNA polymerase. Such an oligonucleotide enables transcription to be initiated at a site of the target which is not normally a transcription start site for the RNA polymerase.

Modified promoter for RNA polymerase, its preparation and its

Oligonucleotide comprising, from 5' to 3', a first portion comprising an upstream segment and a downstream segment forming a stem-loop structure, and a second portion not capable of self-pairing, in which the first portion comprises at least one blocking agent, capable of blocking the replication, by a polymerase, of the said oligonucleotide, such that at least one portion of the downstream segment is replicated and such that the upstream segment is replicated neither totally nor partially. This oligonucleotide can be used as primer in a process for the cyclic amplification of a target sequence of a nucleic acid.

Oligonucleotide which can be used as primer in a method of amplification

Method for nucleic acid amplification by transcription using

A chimera oligonucleotide is provided that can be used in a process for obtaining transcripts and/or amplification of a target sequence of a nucleic acid, having, at its 3' end, a downstream sequence. The oligonucleotide comprises successively, from 5' to 3', 1) a first oligonucleotide segment, of the DNA type, comprising a sense sequence of a promoter of an RNA polymerase, 2) a second oligonucleotide segment, of the DNA type, capable of hybridizing with the downstream sequence, and 3) a third oligonucleotide segment, of the RNA type, capable of hybridizing with a part of the target sequence contiguous to the downstream sequence, the third segment being blocked at 3'. A process using the chimera oligonucleotide and an enzyme system containing DNA polymerase activity, RNA polymerase activity, and a third activity, for example, an RNase H activity provides transcription products of the target. By adding a second chimera oligonucleotide capable of hybridizing with the complement of the target, cyclic amplification of the target and its complement are obtained.

Growing community of inventors

Lyons, France

Pierre Levasseur