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A simple protocol to isolate a single human cell PRDX1 knockout generated by CRISPR-Cas9 system

journal contribution
submitted on 2024-04-03, 10:38 and posted on 2024-04-03, 10:38 authored by Mustapha Aouida, Dina Aljogol, Reem Ali, Dindial Ramotar

Here, we describe a protocol for human PRDX1 gene knockout cells using the CRISPR-Cas9 system. The protocol describes all the steps sequentially: (1) single-guide RNA design, cloning, and transfection; (2) gene editing evaluation by T7EI assay; (3) single-cell isolation; and (4) knockout verification to determine indels in one or both alleles by Sanger sequencing. This strategy is based on the efficiency of DNA editing, avoids antibiotic selection, and bypasses the need for cell sorting.

Other Information

Published in: STAR Protocols
License: http://creativecommons.org/licenses/by-nc-nd/4.0/
See article on publisher's website: https://dx.doi.org/10.1016/j.xpro.2022.101216

History

Language

  • English

Publisher

Elsevier

Publication Year

  • 2022

License statement

This Item is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Institution affiliated with

  • Hamad Bin Khalifa University
  • College of Health and Life Sciences - HBKU

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