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.
Shuldiner
A R
AR
Diabetes Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.
Nirula
A
A
Roth
J
J
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