Establishment of plasma microRNA detection method by using taqman probe based quantitative reverse transcription PCR

MicroRNAs (miRNAs) are a kind of short non—coding RNAs that regulate gene expression at the post—transcriptional level. Recently, many studies have found that circulating miRNAs have the potential to sever as diagnostic biomarkers for many diseases. However, the methods for the quantification of circulating miRNAs still need more adjustment. In this study, we tried to establish a reliable method to quantify the plasma miRNAs. We used quantitative real—time PCR with taqman probes to detect the plasma miR—153 level. Three controls were used in this study, including two external miRNAs control from C. elegans miRNAs (cel—miR—54 and cel—miR—238) and one internal control (hsa—miR—486). All of these controls were stable in plasma and the cel—miR—238/cel—miR—54/hsa—miR—486 combination could improve the normalization process. The expression level of the target miRNA, human plasma miR—153, could be quantified accurately with taqman probes .The assay has high accuracy, high sensitivity and a large dynamic range from 100 copies to 10(13) copies in the PCR reaction. Our study provided a standardized quantification method for plasma miRNAs which might be used as biomarker in many diseases research.