PURPOSE.
We aim to develop better reagents, vectors and methodologies to enable efficient gene silencing, gene activation, and gene editing in cancer cells. This will enable the rapid and precise introduction of defined genetic changes into cancer cells to create the desired genotype for downstream phenotype analysis. This will also enable 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 developed methods for the design and construction of CRISPR/Cas9 libraries. More recently, we have developed reagents and methods for multi-gene targeting using combinatorial siRNA and CRISPR guides. ACCOMPLISHMENT. We have developed reagents and methods for gene targeting in the following areas: 1) customizable CRISPR/Cas9 libraries for gene knockdown; 2) combinatorial gene knockdown using siRNAs; 3) validated siRNA reagents targeting Ras effectors; and 4) combinatorial gene KO using CRISPR guide RNAs.
