1. High-efficiency nonviral CRISPR/Cas9-mediated gene editing of human T cells using plasmid donor DNA.
- Author
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Oh SA, Senger K, Madireddi S, Akhmetzyanova I, Ishizuka IE, Tarighat S, Lo JH, Shaw D, Haley B, and Rutz S
- Subjects
- DNA genetics, Humans, Plasmids genetics, T-Lymphocytes, CRISPR-Cas Systems genetics, Gene Editing methods
- Abstract
Genome engineering of T lymphocytes, the main effectors of antitumor adaptive immune responses, has the potential to uncover unique insights into their functions and enable the development of next-generation adoptive T cell therapies. Viral gene delivery into T cells, which is currently used to generate CAR T cells, has limitations in regard to targeting precision, cargo flexibility, and reagent production. Nonviral methods for effective CRISPR/Cas9-mediated gene knock-out in primary human T cells have been developed, but complementary techniques for nonviral gene knock-in can be cumbersome and inefficient. Here, we report a convenient and scalable nonviral method that allows precise gene edits and transgene integration in primary human T cells, using plasmid donor DNA template and Cas9-RNP. This method is highly efficient for single and multiplex gene manipulation, without compromising T cell function, and is thus valuable for use in basic and translational research., (© 2022 Genentech, Inc.)
- Published
- 2022
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