Isothermal Amplification of RNA by Transcription-Mediated Amplification (TMA)

Abstract

Conventional methods of amplifying an RNA target require a discrete reverse transcriptase step, followed by a separate DNA amplification method (e.g. PCR, LCR, SDA). Here, we describe transcription-mediated amplification (TMA), which amplifies RNA or DNA directly using reverse transcriptase and RNA polymerase (Kacian and Fultz, 1996). TMA has been used to amplify cellular mRNA, viral RNA and highly structured ribosomal RNA (McDonough, Bott and Giachetti, 1997; Jonas et al., 1993). Because RNA is an integral part of its amplification cycle, TMA has excellent sensitivity with these targets - typically <10 copies of mRNA and <50 copies of rRNA. TMA products are single-stranded RNAs, which do not need to be denatured prior to probe detection.