Rapid generation of drug-resistance alleles at endogenous loci using CRISPR-Cas9 indel mutagenesis.

ABSTRACT: Genetic alterations conferring resistance to the effects of chemical inhibitors are valuable tools for validating on-target effects in cells. Unfortunately, for many therapeutic targets such alleles are not available. To address this issue, we evaluated whether CRISPR-Cas9-mediated insertion/deletion (indel) mutagenesis can produce drug-resistance alleles at endogenous loci. This method takes advantage of the heterogeneous in-frame alleles produced following Cas9-mediated DNA cleavage, which we show can generate rare alleles that confer resistance to the growth-arrest caused by chemical inhibitors. We used this approach to identify novel resistance alleles of two lysine methyltransferases, DOT1L and EZH2, which are each essential for the growth of MLL-fusion leukemia cells. We biochemicall

SUBMITTER: Ipsaro JJ 

PROVIDER: S-EPMC5322889 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature