Plasmid- or Ribonucleoprotein-Mediated CRISPR/Cas Gene Editing in Primary Murine T Cells

Methods Mol Biol. 2021:2285:255-264. doi: 10.1007/978-1-0716-1311-5_20.

Abstract

The CRISPR/Cas technology allows for genome editing in primary T cells. We herein describe the activation of primary murine CD4+ or CD8+ T cells, followed by electroporation with plasmid or ribonucleoproteins (RNP) for gene modification. Gene edited T cells can subsequently be transferred to host mice for in vivo studies or cultured in vitro for further characterization. This protocol enables sophisticated genetic analysis of T cells using commonly available virus-free reagents.

Keywords: CRISPR/Cas; Electroporation; Gene editing; Plasmid; Primary murine T cells; RNP.

MeSH terms

  • Animals
  • CD4-Positive T-Lymphocytes / immunology
  • CD4-Positive T-Lymphocytes / metabolism*
  • CD8-Positive T-Lymphocytes / immunology
  • CD8-Positive T-Lymphocytes / metabolism*
  • CRISPR-Cas Systems*
  • Cells, Cultured
  • Electroporation
  • Gene Editing*
  • Lymphocyte Activation
  • Mice
  • Phenotype
  • Plasmids / genetics*
  • Plasmids / metabolism
  • Primary Cell Culture
  • Research Design
  • Ribonucleoproteins / genetics*
  • Ribonucleoproteins / metabolism
  • Workflow

Substances

  • Ribonucleoproteins