Cancer Biology

The expression of genes is frequently manipulated in cell lines to study their cellular functions. The use of exogenous small Interfering RNAs (siRNAs) is a very efficient technique to temporarily downregulate the expression of genes of interest [reviewed by Hannon and Rossi (2004)]. A genome-wide siRNA library allows the user to study both the effect of each individual gene on a particular cell phenotype in a high throughput manner and also assess its phenotypic effect relative to all other genes targeted. Several factors that potentially influence the outcome of a screen need to be considered when performing a large siRNA screen (Jiang et al., 2011). Here we present a detailed protocol for a genome-wide screen to identify genes involved in anti-cancer drug resistance using the human siGENOME library from Dharmacon. In this protocol, we focus on resistance to treatment with the Epidermal Growth Factor Receptor-Tyrosine Kinase Inhibitor (EGFR-TKI) erlotinib in the lung cancer cell line PC9, which is exquisitely sensitive to EGFR-TKIs (de Bruin et al., 2014). This protocol can be used for other cell lines and other drug treatments, as we expand in the Notes below.