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PLoS One DOI:10.1371/journal.pone.0131370

Morphological Profiles of RNAi-Induced Gene Knockdown Are Highly Reproducible but Dominated by Seed Effects.

Publication TypeJournal Article
Year of Publication2015
AuthorsSingh, S, Wu, X, Ljosa, V, Bray, M-A, Piccioni, F, Root, DE, Doench, JG, Boehm, JS, Carpenter, AE
JournalPLoS One
Volume10
Issue7
Pagese0131370
Date Published2015
ISSN1932-6203
KeywordsCell Line, Gene Expression Profiling, Gene Knockdown Techniques, Humans, RNA Interference, RNA, Small Interfering
Abstract

RNA interference and morphological profiling-the measurement of thousands of phenotypes from individual cells by microscopy and image analysis-are a potentially powerful combination. We show that morphological profiles of RNAi-induced knockdown using the Cell Painting assay are in fact highly sensitive and reproducible. However, we find that the magnitude and prevalence of off-target effects via the RNAi seed-based mechanism make morphological profiles of RNAi reagents targeting the same gene look no more similar than reagents targeting different genes. Pairs of RNAi reagents that share the same seed sequence produce image-based profiles that are much more similar to each other than profiles from pairs designed to target the same gene, a phenomenon previously observed in small-scale gene-expression profiling experiments. Various strategies have been used to enrich on-target versus off-target effects in the context of RNAi screening where a narrow set of phenotypes are measured, mostly based on comparing multiple sequences targeting the same gene; however, new approaches will be needed to make RNAi morphological profiling (that is, comparing multi-dimensional phenotypes) viable. We have shared our raw data and computational pipelines to facilitate research.

URLhttp://dx.plos.org/10.1371/journal.pone.0131370
DOI10.1371/journal.pone.0131370
Pubmed

http://www.ncbi.nlm.nih.gov/pubmed/26197079?dopt=Abstract

Alternate JournalPLoS ONE
PubMed ID26197079
PubMed Central IDPMC4511418