Methods and devices for multiplexing amplification reactions

Details Methods and devices for multiplexing amplification reactions Methods and devices for multiplexing amplification reactions

C - Chemistry, Metallurgy

12

Q

C12Q 1/68 (2006.01)

Patent

CA 2410281

The present invention pertains to methods, reagents, compositions, kits and instruments for use in simultaneously amplifying multiple targets. In particular, this invention is based on the discovery of a two-step multiplex amplification reaction wherein the first step truncates the standard initial multiplex amplification roud to "boost" the sample copy number by only a 100 - 1000 fold increase in the target. Following the first step the product is divided into optimized secondary single amplification reactions, each containing one of the primer sets that were used previously in the first or multiplexed booster step. The booster step can occur using an aqueous target nucleic acid or using a solid phase archived nucleic acid.

La présente invention concerne des procédés, des réactifs, des compositions, des nécessaires et des instruments destinés à l'amplification simultanée de cibles multiples. De fait, l'invention repose sur la découverte d'une réaction d'amplification multiplexe en deux temps. Le premier temps correspond à une interruption du cycle standard d'amplification multiplexe initial pour multiplier par 100 à 1000 le nombre d'exemplaires de l'échantillon dans la cible. A la fin de ce premier temps, le produit est divisé donnant autant de réactions d'amplification secondaire optimisées, chacune d'entre elle contenant l'un des jeux d'amorces qui ont servi préalablement pour le premier temps ou pour le temps de multiplication. Ce temps de multiplication peut résulter de l'utilisation d'un acide nucléique en cible aqueuse ou de l'utilisation d'un acide nucléique archivé en phase solide.

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