MicroRNA Isolation from Plasma for Real‐Time qPCR Array. (14th September 2018)

• Record Type: Journal Article
• Authors: Witvrouwen, Isabel; Gevaert, Andreas B.; Van Craenenbroeck, Emeline M.; Van Craenenbroeck, Amaryllis H.
• Editors: Dracopoli, Nicholas C.; Haines, Jonathan L.; Korf, Bruce R.; Morton, Cynthia C.; Seidman, Christine E.; Seidman, J.G.; Smith, Douglas R.
• Is Part Of: Current protocols in human genetics. Volume 99(2018)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2018-09-14
• Subjects: array -- isolation -- microRNA -- plasma; Human genetics -- Laboratory manuals; Genetic Techniques; Human genetics; Laboratory manuals; 599.935028
• DOI: 10.1002/cphg.69
• Languages: English
• ISSNs: 1934-8258

Abstract

Abstract: MicroRNAs are short non‐coding RNAs that regulate gene expression at the post‐transcriptional level by mRNA degradation or suppression of translation. Their stability in plasma makes them attractive biomarkers. Since many plasma microRNA isolation procedures exist and the yield can be highly variable, we recently optimized the microRNA isolation and preamplification procedure using the mir Vana PARIS kit (Thermo Fisher Scientific) for miRNA quantification with TaqMan Low Density Arrays in plasma samples. The method here is slightly modified from the original procedure supplied by Thermo Fisher. Based on our findings, recommendations are the following: (1) use Arabidopsis thaliana (Ath) miR‐159a as spike‐in control, (2) use a 100‐µl elution volume during RNA isolation, and (3) add a preamplification step without dilution of the preamplification product. In this article we provide a step‐by‐step microRNA isolation and quantification procedure using human plasma samples for TaqMan Low Density Arrays. © 2018 by John Wiley & Sons, Inc.