Methods for mammalian genetic analysis

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PURPOSE. We aim to develop better reagents, vector systems and methodologies to enable efficient gene silencing, gene activation, and gene editing in cancer cells. This will enable us to rapidly introduce defined genetic changes into cells to create the desired genotype and study the resulting phenotype. This will also enable us to carry out gain and loss of function genetic screens in cancer cells for target discovery. SIGNIFICANT MATERIALS AND METHODS. We have generated expression vectors for shRNAs, cDNAs and sgRNAs. We have also developed methods for the design and construction of CRISPR/Cas9 libraries. ACCOMPLISHMENT. We have developed reagents and methods for combinatorial gene knockdown and for targeted gene knockout in cancer cells. We have also developed CRISPR/Cas9 libraries that can be used for gene knockout screening in cancer cell lines.