[Adapting Mouse Genome Editing Technique from Scratch Using in utero Electroporation].

Q3 Medicine Molekulyarnaya Biologiya Pub Date : 2024-11-01 DOI:10.31857/S0026898424060142, EDN: IAETMS

J V Popova, V D Bets, E S Omelina, L V Boldyreva, E N Kozhevnikova

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Abstract

Mouse genome modification requires costly equipment and highly skilled personnel to manipulate zygotes. A number of zygote electroporation techniques were reported to be highly efficient in gene delivery. One of these methods called i-GONAD (improved Genome-editing via Oviductal Nucleic Acids Delivery) describes electroporation-based gene transfer to zygotes in utero. Here we adopted this technology to develop an easy-to-use and cost-effective pipeline enabling mouse genome-editing from scratch with minimal requirements to operator skills and animal use. We chose the CRISPR/Cas9 system as a genome editing tool and i-GONAD as a gene delivery method to produce Il10 knockout in C57BL/6 mice. Three animals out of 13 delivered pups (23%) were genetically compromised at Il10 locus suggesting the feasibility of the approach. This protocol provides detailed description of the used technical settings paired with troubleshooting tips and could be of interest to those who aim at establishing in-house mouse transgenesis pipeline at minimal equipment cost from scratch.

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