Generation of scalable cancer models by combining AAV-intron-trap, CRISPR/Cas9, and inducible Cre-recombinase.

ABSTRACT: Scalable isogenic models of cancer-associated mutations are critical to studying dysregulated gene function. Nonsynonymous mutations of splicing factors, which typically affect one allele, are common in many cancers, but paradoxically confer growth disadvantage to cell lines, making their generation and expansion challenging. Here, we combine AAV-intron trap, CRISPR/Cas9, and inducible Cre-recombinase systems to achieve >90% efficiency to introduce the oncogenic K700E mutation in SF3B1, a splicing factor commonly mutated in multiple cancers. The intron-trap design of AAV vector limits editing to one allele. CRISPR/Cas9-induced double stranded DNA breaks direct homologous recombination to the desired genomic locus. Inducible Cre-recombinase allows for the expansion of cells prior to loxp ex

SUBMITTER: Boddu PC 

PROVIDER: S-EPMC8514589 | biostudies-literature | 2021 Oct

REPOSITORIES: biostudies-literature