Reagents and methods for coupled high temperature reverse...

Details Reagents and methods for coupled high temperature reverse... Reagents and methods for coupled high temperature reverse...

C - Chemistry, Metallurgy

12

N

C12N 15/10 (2006.01) C12N 9/12 (2006.01) C12Q 1/68 (2006.01)

Patent

CA 2127188

Methods are provided for the replication and amplification of RNA sequences by thermoactive DNA polymerases. The reverse transcription reaction is performed in an appropriate buffer comprising a metal buffer which buffers the divalent cation concentration and which buffer preferably buffers both the pH and the divalent cation concentration. Said divalent cation is preferably Mn2+. In a preferred embodiment, high temperature reverse transcription is coupled to nucleic acid amplification in a one tube, one enzyme procedure using a thermostable DNA polymerase. Methods for eliminating carry over contamination of amplifications due to prior reverse transcription reactions are also provided. Reagents and kits particularly suited for the methods of the present invention are provided.

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