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. 2022 Jan 17;2:821107. doi: 10.3389/falgy.2021.821107

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Copyright © 2022 Brackett, Pomés and Chapman.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Workflow of experimental approach to delete the Fel d 1 genes using CRISPR-Cas9. Fel d 1 chains 1 and 2 were sequenced to identify conserved regions in the genes to target with CRISPR editing. Guide RNAs (sgRNAs) with sequences complementary to the conserved DNA target regions were designed and synthesized. The Fel d 1-specific sgRNAs and Cas9 nuclease were delivered to immortalized cat cells using lipid-based transfection. Successful in vitro editing, evaluated by DNA sequence decomposition and T7E1 (T7 endonuclease 1) mismatch detection, will guide future in vivo knockouts of Fel d 1.