title: edgeR: a versatile tool for the analysis of shRNA-seq and CRISPR-Cas9 genetic screens.
abstract: Pooled library sequencing screens that perturb gene function in a high-throughput manner are becoming increasingly popular in functional genomics research. Irrespective of the mechanism by which loss of function is achieved, via either RNA interference using short hairpin RNAs (shRNAs) or genetic mutation using single guide RNAs (sgRNAs) with the CRISPR-Cas9 system, there is a need to establish optimal analysis tools to handle such data. Our open-source processing pipeline in edgeR provides a complete analysis solution for screen data, that begins with the raw sequence reads and ends with a ranked list of candidate genes for downstream biological validation. We first summarize the raw data contained in a fastq file into a matrix of counts (samples in the columns, genes in the rows) with options for allowing mismatches and small shifts in sequence position. Diagnostic plots, normalization and differential representation analysis can then be performed using established methods to prioritize results in a statistically rigorous way, with the choice of either the classic exact testing methodology or generalized linear modeling that can handle complex experimental designs. A detailed users' guide that demonstrates how to analyze screen data in edgeR along with a point-and-click implementation of this workflow in Galaxy are also provided. The edgeR package is freely available from http://www.bioconductor.org.
authors: Molecular Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia.
authors: Zhiyin
authors: Z
authors: Dai
authors: Stem Cells and Cancer Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia ; Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia.
authors: Julie M
authors: JM
authors: Sheridan
authors: Molecular Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia ; Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia.
authors: Linden J
authors: LJ
authors: Gearing
authors: Molecular Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia ; Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia.
authors: Darcy L
authors: DL
authors: Moore
authors: Molecular Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia.
authors: Shian
authors: S
authors: Su
authors: Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia ; Systems Biology and Personalised Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia.
authors: Sam
authors: S
authors: Wormald
authors: Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia ; Systems Biology and Personalised Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia.
authors: Stephen
authors: S
authors: Wilcox
authors: Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia ; Systems Biology and Personalised Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia.
authors: Liam
authors: L
authors: O'Connor
authors: Molecular Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia ; Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia.
authors: Ross A
authors: RA
authors: Dickins
authors: Molecular Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia ; Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia.
authors: Marnie E
authors: ME
authors: Blewitt
authors: Molecular Medicine Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, 3052, Australia ; Department of Medical Biology, The University of Melbourne, Parkville, Victoria, 3010, Australia.
authors: Matthew E
authors: ME
authors: Ritchie