Establishment of two homozygous CRISPR interference (CRISPRi) knock-in human induced pluripotent stem cell (hiPSC) lines for titratable endogenous gene repression

Authors

Schoger E, Zimmermann WH, Cyganek L, Zelarayán LC

Journal

Stem Cell Research

Citation

Stem Cell Res. 2021 Jul 27;55:102473.

Abstract

Using nuclease-deficient dead (d)Cas9 without enzymatic activity fused to transcriptional inhibitors (CRISPRi) allows for transcriptional interference and results in a powerful tool for the elucidation of developmental, homeostatic and disease mechanisms. We inserted dCas9KRAB (CRISPRi) cassette into the AAVS1 locus of hiPSC lines, which resulted in homozygous knock-in with an otherwise unaltered genome. Expression of dCas9KRAB protein, pluripotency and the ability to differentiate into all three embryonic germ layers were validated. Furthermore, functional cardiomyocyte generation was tested. The hiPSC-CRISPRi cell lines offer a valuable tool for studying endogenous transcriptional repression with single and multiplexed possibilities in all human cell types.

DOI

10.1016/j.scr.2021.102473

 
Pubmed Link