Focused siRNA screening using validated siRNA libraries

Division Of Basic Sciences - Nci

Investigators

Linked publications, trials & patents

Abstract

BACKGROUNDRNAi has become a powerful genetic tool for dissecting genetic dependencies in cancer cells. Current generation of siRNA libraries suffer from a major disadvantage that there is no validation for the siRNAs in the library and thus the false negative rate is very high siRNA screens.PURPOSEIn this project we aim to accomplish the following goals: 1) constructing a high quality siRNA library targeting Ras family and related genes in which individual siRNAs in the library is knockdown validated; 2) use this siRNA library to screen a panel of human cancer cell lines with or without KRAS mutations to identify those siRNAs that show synthetic lethality among the KRAS mutant cell lines; and 3) functional dissection of the mechanisms of action of candidate synthetic lethal genes using molecular and pharmacological tools.SIGNIFICANT MATERIALS AND METHODSHighly validated siRNA library targeting Ras family and related genes.FY2012 ACCOMPLISHMENTWe have started constructing the siRNA library. We have screened a large number of siRNAs for our genes of interest and identified those that gave potent knockdown. We have tested the efficacy of these siRNAs in a panel of cancer cell lines to demonstrate that they are uniformly potent and can be used for high-throughput siRNA screens.