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Design, Manufacture, and Assay of the Efficacy of siRNAs for Gene Silencing

  • Louise A. Dawson
  • Badar A. Usmani
Part of the Methods in Molecular Biology™ book series (MIMB, volume 439)

Abstract

Small interfering RNAs (siRNAs) have been widely exploited for nucleotide-sequence-specific posttranscriptional gene silencing, as a tool to investigate gene function in eukaryotes, and they hold promise as potential therapeutic agents. Conventionally designed siRNAs are 21-mers with symmetric 2-nt 3′ overhangs that mimic intermediates (microRNAs or miRNAs) of the natural processing of longer dsRNA (double-stranded RNA). siRNAs are sequences with full c omplementarity to their target mRNA and can be generated by either chemical synthesis or processing of shRNAs (short hairpin RNAs) transcribed from DNA vectors. To minimize off-target effects, any homology to nontarget mRNA can be verified using the expressed sequence tag (EST) database for the relevant organism. Here, we provide a practical guide and an overview to the design and selection of effective and specifc siRNAs.

Keywords

small interfering RNA gene silencing endothelin-converting enzyme prostate cancer invasion stromal-epithelial interactions 

Notes

Acknowledgments

This work was supported by Yorkshire Cancer Research (YCR) and Prostate Cancer Research Foundation (PCRF).

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Louise A. Dawson
    • 1
  • Badar A. Usmani
    • 1
  1. 1.Institute of Cellular and Molecular BiologyUniversity of LeedsLeedsUK

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