Robust CRISPR/Cas9-Mediated Tissue-Specific Mutagenesis Reveals Gene Redundancy and Perdurance in Drosophila.

ABSTRACT: Tissue-specific loss-of-function (LOF) analysis is essential for characterizing gene function. Here, we present a simple, yet highly efficient, clustered regularly interspaced short palindromic repeats (CRISPR)-mediated tissue-restricted mutagenesis (CRISPR-TRiM) method for ablating gene function in Drosophila This binary system consists of a tissue-specific Cas9 and a ubiquitously expressed multi-guide RNA (gRNA) transgene. We describe convenient toolkits for making enhancer-driven Cas9 lines and multi-gRNAs that are optimized for mutagenizing somatic cells. We demonstrate that insertions or deletions in coding sequences more reliably cause somatic mutations than DNA excisions induced by two gRNAs. We further show that enhancer-driven Cas9 is less cytotoxic yet results in more comp

SUBMITTER: Poe AR 

PROVIDER: S-EPMC6366929 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature