Application of Oligo-DT Molecules To Avoid Generation of High Molecular PCR Products Induced By Poly-A Carrier

摘要

During RNA isolation carrier nucleic acids such as polyA-RNA are used in order to increase the yield of the isolated RNA. The carrier nucleic acids may lead to high molecular weight products which may interfere in subsequent steps, such as reverse transcription, PCR and gel electrophoresis. The present invention therefore refers to a method and a kit for reverse transcription and the use of a blocking nucleic acid molecule for blocking the carrier polyA-RNA.

主权项

1. A method of reverse transcription, wherein the method comprises the steps of
a) providing a sample comprising RNA, b) contacting the sample with a carrier nucleic acid thereby generating a mixture, c) applying the mixture to a matrix under conditions such that binding of the RNA and the carrier nucleic acid to the matrix occurs, d) separating the matrix with the RNA and the carrier nucleic acid bound to the matrix from the mixture, e) eluting the RNA and the carrier nucleic acid from the matrix thereby generating an eluate, f) adding a blocking nucleic acid to the eluate under conditions such that hybridization of the blocking nucleic acid to the carrier nucleic acid occurs, g) adding a primer to the eluate from step f) under conditions such that hybridization of the primer to the RNA occurs, and h) reverse transcribing the RNA into cDNA.