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Nat Protoc. 2014 Oct;9(10):2341-53. doi: 10.1038/nprot.2014.160. Epub 2014 Sep 11.

Measuring genetic interactions in human cells by RNAi and imaging.

Author information

1
1] German Cancer Research Center (DKFZ), Division of Signaling and Functional Genomics, Heidelberg, Germany. [2] Department of Cell and Molecular Biology, Heidelberg University, Heidelberg, Germany.
2
European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany.

Abstract

Observation of how genetic interactions modulate phenotypes is a powerful method for dissecting their underlying molecular and functional networks. Whereas in model organisms genetic interaction studies are well established, systematic analysis of genetic interactions in human cells has remained challenging. Here we provide a detailed protocol for large-scale mapping of genetic interactions in human cells using a high-throughput phenotyping approach. Pairwise gene product depletion is induced by siRNA-mediated knockdown, and the resulting phenotypes are quantified by automated imaging and computational analysis to provide the basis for detecting genetic interactions between all pairs of genes tested. The whole workflow, depending on the size of the experiment, takes 3 or more weeks to complete.

PMID:
25211512
DOI:
10.1038/nprot.2014.160
[Indexed for MEDLINE]

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