Linked linear amplification of nucleic acids

C - Chemistry – Metallurgy – 12 – Q

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C12Q 1/68 (2006.01) C12N 15/10 (2006.01) C12P 19/34 (2006.01)

Patent

CA 2167840

The extensive synthesis ("amplification") of a nucleic acid sequence of interest is attained through a linked series of multi-cycle primer extension reactions (LLA). The primers used in each of the primer extension reactions of the process contain non-replicable elements that halt nucleic acid synthesis and thereby prevent the synthesis molecules from serving as templates in subsequent cycles. Synthesized molecules accumulate during primer extension in a mathematically linear fashion, thereby rendering the process relatively insensitive to contaminating nucleic acids. Multiple primer sets are employed, thereby ensuring the accumulation of a large number of copies of the nucleic acid sequence of interest. The invention also provides for the detection of an amplified nucleic acid sequence of interest, as well as reagent kits for carrying out the reaction.

On effectue la synthèse extensive ("amplification") d'une séquence d'acide nucléique à étudier au moyen d'une série enchaînée de réactions d'extension d'amorces à plusieurs cycles. Les amorces utilisées dans chacune des réactions d'extension d'amorce du procédé contiennent des éléments non réplicables qui stoppent la synthèse d'acide nucléique et empêchent par conséquent les molécules synthétisées de servir de matrice dans les cycles subséquents. Les molécules synthétisées s'accumulent de manière mathématiquement linéaire pendant l'extension de l'amorce et confèrent au procédé une relative insensibilité par rapport aux acides nucléiques contaminants. Plusieurs ensembles d'amorces sont utilisés afin d'assurer l'accumulation d'un grand nombre de copies de la séquence d'acide nucléique à étudier. L'invention concerne également la détection d'une séquence d'acide nucléique à étudier ainsi que des trousses de réactifs permettant d'effectuer une telle réaction.

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