Brief Communication abstract
Nature Methods 6, 443 - 445 (2009)
Published online: 17 May 2009 | doi:10.1038/nmeth.1330
Rapid creation and quantitative monitoring of high coverage shRNA libraries
Michael C Bassik1,4, Robert Jan Lebbink2,4, L Stirling Churchman1, Nicholas T Ingolia1, Weronika Patena1,2, Emily M LeProust3, Maya Schuldiner1, Jonathan S Weissman1 & Michael T McManus2
Short hairpin RNA libraries are limited by low efficacy of many shRNAs and by off-target effects, which give rise to false negatives and false positives, respectively. Here we present a strategy for rapidly creating expanded shRNA pools (
30 shRNAs per gene) that are analyzed by deep sequencing (EXPAND). This approach enables identification of multiple effective target-specific shRNAs from a complex pool, allowing a rigorous statistical evaluation of true hits.
- Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biomedical Research, and Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, California, USA.
- Department of Microbiology and Immunology, and University of California San Francisco Diabetes Center, University of California, San Francisco, San Francisco, California, USA.
- Genomics Solution Unit, Agilent Technologies Inc., Santa Clara, California, USA.
- These authors contributed equally to this work.
Correspondence to: Jonathan S Weissman1 e-mail: weissman@cmp.ucsf.edu
Correspondence to: Michael T McManus2 e-mail: michael.mcmanus@ucsf.edu
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