CRISPRi-mediated transcriptional silencing in iPSCs for the study of human brain development

Pia Annette Johansson, Anita Adami, Johan Jakobsson.

STAR Protocols, 2022

This protocol describes using CRISPRi-mediated transcriptional silencing in human induced pluripotent stem cells. In addition, it contains an efficient protocol for neural progenitor differentiation. The method is directly applicable to loss-of-function studies in brain development research. Biolaminin 521 (laminin-521) is applied as the cell culture matrix for PSCs and Biolaminin 111 (laminin-111) for hiPSC differentiation into forebrain neural progenitor cells. The protocol is expected to achieve high transduction and silencing efficiency. After two weeks of differentiation, the authors detected no reduction in the percentage of positive cells, and the cerebral organoids had stable transcriptional silencing even after 4 months.