Method for unbiased mrna amplification

C - Chemistry – Metallurgy – 12 – Q

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C12Q 1/68 (2006.01)

Patent

CA 2311238

Methods of enzymatically producing unbiased amounts of nucleic acid from mRNA are provided. In the subject methods, a sample of mRNA is converted to ds cDNA using a primer containing an RNA polymerase site and at least one priming site. The resultant ds cDNA is then asymmetrically amplified with captureable primer to produce captureable cDNA which is then converted to captured ds cDNA. The resultant ds cDNA may then be used in a number of different applications, such as in the preparation of amplified amounts of aRNA, in the preparation of cDNA probes, and the like.

L'invention concerne des techniques enzymatiques qui permettent de produire des quantités non biaisées d'acide nucléique à partir d'ARNm. Selon ces techniques, on convertit un échantillon d'ARNm en ADNc double brin en utilisant une amorce qui contient un site promoteur d'ARN polymérase et au moins un site d'amorçage. Puis on amplifie asymétriquement l'ADNc double brin obtenu avec une amorce capturable, de façon à produire un ADNc capturable, que l'on convertit ensuite en ADNc double brin capturé. On peut utiliser l'ADNc double brin obtenu dans de nombreuses applications, telles que la préparation de quantités amplifiées d'ARNa, la préparation de sondes d'ADNc, etc.

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