Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays
2019
Gene perturbation studies have been extensively used to investigate the role of individual genes in AML pathogenesis. For achieving complete gene disruption, many of these studies have made use of complex
gene knockoutmodels. While these studies with knockout mice offer an elegant and time-tested system for investigating genotype-to-phenotype relationships, a rapid and scalable method for assessing
candidate genesthat play a role in AML cell proliferation or survival in AML models will help accelerate the parallel interrogation of multiple
candidate genes. Recent advances in
genome-editingtechnologies have dramatically enhanced our ability to perform genetic perturbations at an unprecedented scale. One such system of
genome editingis the
CRISPR-
Cas9-based method that can be used to make rapid and efficacious alterations in the target cell genome. The ease and scalability of
CRISPR/
Cas9-mediated gene-deletion makes it one of the most attractive techniques for the interrogation of a large number of genes in phenotypic assays. Here, we present a simple assay using
CRISPR/
Cas9mediated gene-disruption combined with high-throughput flow-cytometry-based competition assays to investigate the role of genes that may play an important role in the proliferation or survival of human and murine AML cell lines.
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