Generation of Human Isogenic Induced Pluripotent Stem Cell Lines with CRISPR Prime Editing.

ABSTRACT: We developed an efficient CRISPR prime editing protocol and generated isogenic-induced pluripotent stem cell (iPSC) lines carrying heterozygous or homozygous alleles for putatively causal single nucleotide variants at six type 2 diabetes loci (ABCC8, MTNR1B, TCF7L2, HNF4A, CAMK1D, and GCK). Our two-step sequence-based approach to first identify transfected cell pools with the highest fraction of edited cells significantly reduced the downstream efforts to isolate single clones of edited cells. We found that prime editing can make targeted genetic changes in iPSC and optimization of system components and guide RNA designs that were critical to achieve acceptable efficiency. Systems utilizing PEmax, epegRNA modifications, and MLH1dn provided signific

SUBMITTER: Bonnycastle LL 

PROVIDER: S-EPMC10880268 | biostudies-literature | 2024 Feb

REPOSITORIES: biostudies-literature