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Epigenetic silencing is a barrier to editing the X chromosome using the SEC-based CRISPR/Cas9 knock-in protocol in C. elegans

Year: 2025

Type: article

Abstract: . Here, we report a lack of success targeting the X chromosome using this method. CRISPR/Cas9 works as intended, but subsequent floxing of the SEC is blocked. Given that the X chromosome is epigenetically silenced in primordial germ cells (PGCs), this is a logical result. To circumvent this barrier, we suppressed polycomb repressive complex 2 (PRC2) with RNAi to transiently and reversibly reduce silencing in the PGCs, creating a brief window where the X chromosome is amenable to floxing without compromising germ line development. Overall, our results reveal a previously unrecognized limitation of SEC-based CRISPR/Cas9 knock-in and identify a reliable workaround for tagging proteins encoded on the X chromosome.

Source: PubMed

DOI: 10.17912/micropub.biology.001974

Authors: Ryka Iyer, Simon Ferreria, Laahya Guvvala, Hugh Borden, Aariv Arora, Rebecca Shirsat, Ryan Doonan

Language: English

Topic: CRISPR and Genetic Engineering

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: green

OpenAlex ID: W7125257891

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