Protocol of CRISPR-Cas9 knockout screens for identifying ferroptosis regulators

  • STAR Protoc. 2023 Dec 3;4(4):102762. doi: 10.1016/j.xpro.2023.102762.
Xin Yang  1 ,  Shoufu Duan  1 ,  Zhiming Li  1 ,  Zhe Wang  1 ,  Ning Kon  1 ,  Zhiguo Zhang  2 ,  Wei Gu  3
Affiliations
  • 1. Institute for Cancer Genetics, and Herbert Irving Comprehensive Cancer Center, Vagelos College of Physicians & Surgeons, Columbia University, New York, NY, USA.
  • 2. Institute for Cancer Genetics, and Herbert Irving Comprehensive Cancer Center, Vagelos College of Physicians & Surgeons, Columbia University, New York, NY, USA; Department of Pediatrics, and Department of Genetics and Development, Vagelos College of Physicians & Surgeons, Columbia University, New York, NY, USA. Electronic address: [email protected].
  • 3. Institute for Cancer Genetics, and Herbert Irving Comprehensive Cancer Center, Vagelos College of Physicians & Surgeons, Columbia University, New York, NY, USA; Department of Pathology and Cell Biology, Vagelos College of Physicians & Surgeons, Columbia University, New York, NY, USA. Electronic address: [email protected].
PMID: 38048220 DOI: 10.1016/j.xpro.2023.102762
Abstract

Ferroptosis, an iron-dependent programmed cell death triggered by excessive lipid peroxidation, has shown promising therapeutic potentials in human diseases. Here, we describe a protocol of a CRISPR-Cas9 loss-of-function screen to identify regulators in response to different inducers of Ferroptosis. We emphasize the steps of library amplification, drug treatment, high-throughput Sequencing preparation, and bioinformatics analysis using model-based analysis of genome-wide CRISPR-Cas9 knockout (MAGeCK). We also present a method to discover the regulators of Ferroptosis and verify the potential targets efficiently. For complete details on use and execution of this protocol, please refer to Yang et al. (2023).1.

Keywords
Bioinformatics; CRISPR; Cell Biology; Cell culture; Cell-based Assays; Molecular Biology; Sequence analysis; Sequencing.
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