Methods and compositions for the detection and analysis of...

C - Chemistry – Metallurgy – 12 – Q

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C12Q 1/68 (2006.01) C12Q 1/00 (2006.01)

Patent

CA 2598096

The present invention relates to a novel integrated PCR-free signal amplification polynucleotide detection system which combines a specific receptor, an optical transducer, and an amplification mechanism. This novel detection system is based on different electrostatic interactions and conformations between a cationic polythiophene (i.e. polymer 1) and single- stranded or double-stranded polynucleotides (such as ss-DNA or ds-DNA), and the efficient energy transfer between the triplex (complexation between the cationic polythiophene and ds-DNA) and neighboring fluorophores attached to ss- DNA or ds-DNA probes. It is to be understood that in the case of ss-DNA, triplex formation occurs via the hybridization of complementary ss-DNA strands, combined with complexation with the cationic polythiophene. The present detection system allows for the detection of single nucleotide polymorphisms (SNPs) from samples in only a few minutes, without the need for nucleic acid amplification.

Cette invention se rapporte à un nouveau système intégré de détection de polynucléotides par amplification du signal, sans PCR, combinant un récepteur spécifique, un transducteur optique, et un mécanisme d'amplification. Ce nouveau système de détection repose sur différentes interactions électrostatiques et conformations entre un polythiophène cationique (polymère 1) et des polynucléotides monocaténaires ou bicaténaires (tels qu'ADNss ou ADNds), et le transfert d'énergie efficace entre le triplex (complexation du polythiophène cationique avec l'ADNds) et les fluorophores contigus liés aux sondes d'ADNss ou d'ADNds. Il est à noter que dans le cas de l'ADNss, la formation du triplex est réalisée par hybridation d'un brin d'ADNss complémentaire, combinée à la complexation avec le polythiophène cationique. Le système de détection décrit permet la détection de polymorphismes mononucléotidiques (SNP) dans des échantillons en quelques minutes seulement, sans nécessiter une amplification de l'acide nucléique.

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