Targeted Deletion of Vitamin D receptor Gene in Mammalian Cells by CRISPR/Cas9 Systems

BioRxiv : the Preprint Server for Biology
Tao ZhangZhiying Zhang

Abstract

CRISPR/Cas9 system has become a new versatile technology for genome engineering. It utilizes a single guide RNA (sgRNA) to recognize target sequences in genome function, and activates Cas9 endonucleases to cut the locus. In this study, we designed two target sites from conserved regions of vitamin D receptor (VDR) gene in mammalian cells, which cover more than 17 kb of chromosome region depending on the species. The efficacy of single sgRNA mediated gene specific modification was about 22% to 36%. Concurrently, targeted deletions of the intervening genomic segments were generated in chromosomes when the two sgRNAs worked simultaneously. The large genomic DNA segments ranging from 17.8Kb to 23.4 Kb could be precisely deleted in human and mouse chromosomes. Furthermore, the expression level of 24-hydroxylase (CYP24A1) regulated by VDR was significantly increased in cells treated with VDR CRISPR/Cas9 vectors. This study showed that CRISPR/Cas9 system can be employed to generate large genomic segment deletions in different species, providing sgRNAs are designed within conserved regions.

Related Concepts

Chromosomes
Gene Deletion
Genes
Genetic Engineering
Genome
RNA
Tyrosine 3-Monooxygenase
RNA, Guide
Vitamin D3 Receptor
Site

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