A protocol for karyotyping and genetic editing of induced pluripotent stem cells with homology-directed repair mediated CRISPR/Cas9.

ABSTRACT: CRISPR/Cas9-mediated homology-directed repair (HDR) allows precise gene editing, but its efficiency remains low for certain cell types, such as human induced pluripotent stem cells (hiPSCs). In this study, we aimed to introduce the CTNNA1: c.2023C>T (p.Q675*) genetic alteration, which is associated with Hereditary Diffuse Gastric Cancer, into hiPSCs using CRISPR/Cas9. We designed a single-guide RNA targeting the alteration site and a single-stranded oligonucleotide donor DNA template for HDR-based repair. Herein, we report the successful introduction of the CTNNA1: c.2023C>T homozygous alteration in one hiPSC line, which resulted in severe phenotypic changes, including impaired colony formation and cell proliferation. Additionally, we established a straightforward protocol to

SUBMITTER: Lobo S 

PROVIDER: S-EPMC11930342 | biostudies-literature | 2025

REPOSITORIES: biostudies-literature