Journal article

Efficient CRISPR/Cas9 gene ablation in uncultured naïve mouse T cells for in vivo studies

Simone Nüssing, Imran House, Conor Kearney, Stephin Vervoort, Paul Beavis, Jane Oliaro, Ricky Johnstone, Joseph Trapani, Ian Parish

Cold Spring Harbor Laboratory | Published : 2019


CRISPR/Cas9 technologies have revolutionised our understanding of gene function in complex biological settings, including T cell immunology. Current CRISPR-mediated gene deletion strategies in T cells require in vitro stimulation or culture that can both preclude studies of gene function within unmanipulated naïve T cells and can alter subsequent differentiation. Here we demonstrate highly efficient gene deletion within uncultured primary naïve murine CD8 + T cells by electroporation of recombinant Cas9/sgRNA ribonucleoprotein immediately prior to in vivo adoptive transfer. Using this approach, we generated single and double gene knock-out cells within multiple mouse infection models. Striki..

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