Reduced inhibition of one-step rt-pcr

C - Chemistry – Metallurgy – 12 – P

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C12P 19/34 (2006.01) C12Q 1/68 (2006.01) C12N 9/12 (2006.01)

Patent

CA 2706444

The present invention provides a method for amplifying a nucleic acid molecule. The method involves mixing an RNA template with a composition having a reverse transcriptase, a DNA polymerase and a RT inhibition reducer. The RT inhibition reducer can be Sso7d, Sac7d, Sac7e, Sso7e, AluI methylase, suramin, a phosphorothioate oligodeoxycytosine, a phosphorothioate oligodeoxyadenine, a phosphorothioate oligodeoxythymine or poly(rA)(dT). The mixing forms a mixture that is incubated under conditions sufficient to synthesize a DNA molecule complementary to all or a portion of the RNA template, thereby amplifying the nucleic acid molecule.

L'invention concerne un procédé pour amplifier une molécule d'acide nucléique. Le procédé implique le mélange d'une matrice d'ARN avec une composition ayant une transcriptase inverse, un ADN polymérase et un réducteur d'inhibition de RT. Le réducteur d'inhibition de RT peut être de la méthylase de Sso7d, Sac7d, Sac7e, Sso7e, AluI, de la suramine, une oligodéoxycytosine de phosphorothioate, une oligodeoxyadénine de phosphorothioate, une oligodeoxythymine de phosphorothioate ou du poly(Ra) (dT). L'étape de mélange forme un mélange qui est incubé dans des conditions suffisantes pour synthétiser une molécule d'ADN complémentaire à la totalité ou à une partie de la matrice d'ARN, en amplifiant de plus la molécule d'acide nucléique.

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