High-Throughput RNAi Screening for the Identification of Novel Targets

  • Meredith C. Henderson
  • David O. Azorsa
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 986)

Abstract

Gene silencing through RNA interference has provided researchers with an effective way to study gene function. High-throughput RNA interference (HT-RNAi) screening has further permitted researchers to identify functionally relevant mediators of cellular response on a large scale. These screens have greatly expedited the discovery of novel targets and pathway mediators. Here, we describe the methodology for performing HT-RNAi screening of HeLa cells transfected with short interfering RNA (siRNA) libraries in 384-well microplate format. Using this plate format, the HT-RNAi assay can be easily adapted to semi-automated or fully automated platforms. The library siRNA are introduced into the cells through reverse transfection using cationic lipids. HT-RNAi screening for modulators of cell proliferation can be accomplished using a single read out reagent. This type of RNAi screening can be used with most plate-based cellular assays and can be optimized for most cultured cells lines, thus becoming a powerful tool to identify specific gene modulators and targets for drug discovery.

Key words

High-throughput RNAi siRNA Library Assay Screening 

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Copyright information

© SpringerScience+Business Media New York 2013

Authors and Affiliations

  • Meredith C. Henderson
    • 1
  • David O. Azorsa
    • 1
  1. 1.Clinical Translational Research DivisionTranslational Genomics Research Institute (TGen)ScottsdaleUSA

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