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miRNA Expression Profiling Using Illumina Universal BeadChips

  • Jing Chen
  • Craig S. April
  • Jian-Bing FanEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 822)

Abstract

We have developed a highly sensitive, specific, and reproducible method for microRNA (miRNA) expression profiling, using BeadArray technology. This method incorporates an enzyme-assisted specificity step, a solid-phase primer extension to distinguish between members of miRNA families. In addition, a universal PCR is used to amplify all targets prior to array hybridization. Using this method, highly reproducible miRNA expression profiles were generated with 100–200 ng total RNA input. The method has a 3.5–4 log (105–109 molecules) dynamic range and is able to detect 1.2- to 1.3-fold differences between samples. Expression profiles generated by this method are highly comparable to those obtained with RT-PCR (R 2 = 0.85–0.90) and direct sequencing (R = 0.87–0.89). This method should prove useful for high-throughput expression profiling of miRNAs in large numbers of tissue samples.

Key words

miRNA Gene expression analysis Microarray BeadArray DASL assay 

Notes

Acknowledgments

We would like to thank Shawn Baker, Tanya Boyaniwsky, Kirt Haden, Mark Staebell, Christopher Streck, Scott Taylor, Joanne Yeakley, and John Stuelpnagel who are or were at Illumina; Louise Laurent, and Jeanne Loring at The Scripps Research Institute; Renee Rubio, Kristina Holton, and John Quackenbush at Dana-Farber Cancer Institute; Hua Gu at Columbia University and Guoping Fan at UCLA, for helpful discussions.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Illumina, Inc.San DiegoUSA

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