Identification of Genes Regulating Hepatocyte Injury by a Genome-Wide CRISPR-Cas9 Screen.
Document Type
Article
Publication Date
2022
Identifier
DOI: 10.1007/978-1-0716-2557-6_17
Abstract
Gene editing introduces stable mutations into the genome and has powerful applications extending from research to clinical gene therapy. CRISPR-Cas9 gene editing can be employed to study directly the functional impact of stable gene knockout, activation, and knockdown. Here, we describe the end-to-end methodology by which we employ genome-wide CRISPR-Cas9 knockout to study drug toxicity using acetaminophen (APAP) in a hepatocellular carcinoma liver model as an example. This methodology can be extended to other proliferative cell types and chemical metabolic and toxicity models. By employing a massively parallelized genome-wide knockout model, the genes responsible for cellular toxicity and proliferation may be assayed concurrently. Resultant data are interrogated in the context of existing gene expression data, pathway analysis, drug-gene interactions, and orthogonal confirmatory assays to better understand the metabolic mechanisms.
Journal Title
Methods in molecular biology (Clifton, N.J.)
Volume
2544
First Page
227
Last Page
251
MeSH Keywords
Acetaminophen; CRISPR-Cas Systems; Gene Editing; Genome; Hepatocytes
Keywords
CRISPR-Cas9; Drug screen; Gene-editing; Genome-wide; Hepatocytes; Next-generation sequencing; Toxicity
Recommended Citation
Shortt K, Heruth DP. Identification of Genes Regulating Hepatocyte Injury by a Genome-Wide CRISPR-Cas9 Screen. Methods Mol Biol. 2022;2544:227-251. doi:10.1007/978-1-0716-2557-6_17
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