RNA template-specific polymerase chain reaction (RS-PCR): a novel strategy to reduce dramatically false positives.
We report a novel modification of the reverse transcription-polymerase chain reaction method that we have dubbed RNA template-specific PCR (RS-PCR). With this approach, the 5' end of the first strand is tagged with a unique nucleotide sequence during reverse transcription which may then be exploited to amplify preferentially RNA-derived sequences. In our hands, RS-PCR greatly reduces the frequency of false positives and virtually eliminates carryover contamination from DNA fragments amplified in previous experiments.
Diabetes Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.
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Shuldiner
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Nirula
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Roth
D000818:Animals; D001483:Base Sequence; D007328:Insulin / Q000235:genetics; D007515:Islets of Langerhans / Q000378:metabolism; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction / Q000379:methods*; D012313:RNA / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D013698:Templates, Genetic; D014981:Xenopus
D007328:Insulin; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D012313:RNA