Purification of specific DNA species using the CRISPR system

Toshitsugu Fujita; Hodaka Fujii

doi:10.1093/biomethods/bpz008

Purification of specific DNA species using the CRISPR system

Biol Methods Protoc. 2019 Jul 8;4(1):bpz008. doi: 10.1093/biomethods/bpz008. eCollection 2019.

Authors

Toshitsugu Fujita¹, Hodaka Fujii¹

Affiliation

¹ Department of Biochemistry and Genome Biology, Hirosaki University Graduate School of Medicine, Zaifu-cho 5, Hirosaki, Aomori, Japan.

Abstract

In 2013, we developed a new method of engineered DNA-binding molecule-mediated chromatin immunoprecipitation that incorporates the clustered regularly interspaced short palindromic repeats (CRISPR) system to purify specific DNA species. This CRISPR-mediated purification can be performed in-cell or in vitro; CRISPR complexes can be expressed to tag target DNA sequences in the cells to be analyzed, or a CRISPR ribonucleoprotein complex consisting of recombinant nuclease-dead Cas9 (dCas9) and synthetic guide RNA can be used to tag target DNA sequences in vitro. Both methods enable purification of specific DNA sequences in chromatin structures for subsequent identification of molecules (proteins, RNAs, and other genomic regions) associated with the target sequences. The in vitro method also enables enrichment of purified DNA sequences from a pool of heterogeneous sequences for next-generation sequencing or other applications. In this review, we outline the principle of CRISPR-mediated purification of specific DNA species and discuss recent advances in the technology.

Keywords: CRISPR; DNA purification; NGS; dCas9; enChIP.

Publication types

Review