One-step real time RT-PCR for detection of microRNAs

Rapid and simple methods for microRNA (miRNA) detection are essential for biological research of miRNAs and clinical diagnosis. Here we describe a sensitive and specific real time RT-PCR (also RT-qPCR) method for miRNA quantification. The whole detection process including reverse transcription and PCR is performed in one PCR tube by a one-step operation on a real-time PCR system. The results display a wide linear range from 0.1 amol to 10 fmol with a detection limit of 12.6 zmol for miRNA let-7a detection. Let-7a in small RNA samples extracted from tumor cells has been successfully detected by this method. This method is cost-effective, simple and rapid, and has the advantages in the high-throughput routing assay of given miRNAs, as well as in non-model research that has less specific kits and reagents.