High-throughput RNA interference screening using pooled shRNA libraries and next generation sequencing
The Breakthrough Breast Cancer Research Centre, The Institute of Cancer Research, 237 Fulham Road, London, SW3 6JB, UK
Genome Biology 2011, 12:R104 doi:10.1186/gb-2011-12-10-r104Published: 21 October 2011
RNA interference (RNAi) screening is a state-of-the-art technology that enables the dissection of biological processes and disease-related phenotypes. The commercial availability of genome-wide, short hairpin RNA (shRNA) libraries has fueled interest in this area but the generation and analysis of these complex data remain a challenge. Here, we describe complete experimental protocols and novel open source computational methodologies, shALIGN and shRNAseq, that allow RNAi screens to be rapidly deconvoluted using next generation sequencing. Our computational pipeline offers efficient screen analysis and the flexibility and scalability to quickly incorporate future developments in shRNA library technology.