Combined RT-qPCR of mRNA and microRNA Targets within One Fluidigm Integrated Fluidic Circuit

J Biomol Tech. 2016 Jul;27(2):75-83. doi: 10.7171/jbt.16-2702-003. Epub 2016 Mar 7.

Abstract

The ability to profile expression levels of a large number of mRNAs and microRNAs (miRNAs) within the same sample, using a single assay method, would facilitate investigations of miRNA effects on mRNA abundance and streamline biomarker screening across multiple RNA classes. A protocol is described for reverse transcription of long RNA and miRNA targets, followed by preassay amplification of the pooled cDNAs and quantitative PCR (qPCR) detection for a mixed panel of candidate RNA biomarkers. The method provides flexibility for designing custom target panels, is robust over a range of input RNA amounts, and demonstrated a high assay success rate.

Keywords: high-throughput quantification; long/short RNA.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, N.I.H., Extramural

MeSH terms

  • Adult
  • Biomarkers / blood
  • Case-Control Studies
  • Gene Expression Profiling / instrumentation
  • Gene Expression Profiling / methods*
  • Humans
  • MicroRNAs / blood
  • MicroRNAs / genetics*
  • Microfluidic Analytical Techniques
  • RNA, Messenger / blood
  • RNA, Messenger / genetics*
  • Reverse Transcriptase Polymerase Chain Reaction

Substances

  • Biomarkers
  • MicroRNAs
  • RNA, Messenger