Poster Presentation 39th Annual Lorne Genome Conference 2018

Overcoming off target effects in CRISPR and RNAi loss of function screens (#237)

Joseph Rosenbluh 1
  1. Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia

Loss of function screens provide the means to modulate gene expression and permit to systematically study gene function. By comparing RNAi and CRISPR we found that seed mediated RNAi off target effects complicate the interpretation of RNAi based approaches and that CRISPR provides a highly specific approach to modulate gene expression. However, DNA amplifications which are common in many cancer cell lines mediate a Cas9 induced DNA damage response that results in a gene independent proliferation arrest. To overcome this hurdle, we have adopted CRISPR interference (CRISPRi) as an additional approach to suppress gene expression. We found that CRISPRi is not effected by DNA amplifications however, bidirectional promoters (present in 10% of human genes) score as false positives in CRISPRi screens. We demonstrate the utility of combining these approaches as a highly specific approach to study how genes are regulated in normal cells and how they are deregulated during disease progression.