Abstract
Cis-regulatory elements (CREs) play a pivotal role in spatiotemporal control of tissue-specific gene expression, yet the molecular composition of the vast majority of CREs in native chromatin remains unknown. In this article, we describe the clustered regularly interspaced short palindromic repeats (CRISPR) affinity purification in situ of regulatory elements (CAPTURE) approach to simultaneously identify locus-specific chromatin-regulating protein complexes and long-range DNA interactions. Using an in vivo biotinylated nuclease-deficient Cas9 (dCas9) protein and programmable single guide RNAs (sgRNAs), this approach allows for high-resolution and locus-specific isolation of protein complexes and long-range chromatin looping associated with single copy CREs in mammalian cells. Unbiased analysis of the compositional structure of developmentally regulated or disease-associated CREs identifies new features of transcriptional regulation. Hence, CAPTURE provides a versatile platform to study genomic locus-regulating chromatin composition in a mammalian genome.
| Original language | English (US) |
|---|---|
| Article number | e64 |
| Journal | Current Protocols in Molecular Biology |
| Volume | 123 |
| Issue number | 1 |
| DOIs | |
| State | Published - Jul 1 2018 |
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Keywords
- chromatin
- cis-regulatory elements
- CRISPR
- DNA looping
- enhancer
ASJC Scopus subject areas
- Molecular Biology
Cite this
CAPTURE : In Situ Analysis of Chromatin Composition of Endogenous Genomic Loci by Biotinylated dCas9. / Liu, Xin; Zhang, Yuannyu; Chen, Yong; Li, Mushan; Shao, Zhen; Zhang, Michael Q.; Xu, Jian.
In: Current Protocols in Molecular Biology, Vol. 123, No. 1, e64, 01.07.2018.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - CAPTURE
T2 - In Situ Analysis of Chromatin Composition of Endogenous Genomic Loci by Biotinylated dCas9
AU - Liu, Xin
AU - Zhang, Yuannyu
AU - Chen, Yong
AU - Li, Mushan
AU - Shao, Zhen
AU - Zhang, Michael Q.
AU - Xu, Jian
PY - 2018/7/1
Y1 - 2018/7/1
N2 - Cis-regulatory elements (CREs) play a pivotal role in spatiotemporal control of tissue-specific gene expression, yet the molecular composition of the vast majority of CREs in native chromatin remains unknown. In this article, we describe the clustered regularly interspaced short palindromic repeats (CRISPR) affinity purification in situ of regulatory elements (CAPTURE) approach to simultaneously identify locus-specific chromatin-regulating protein complexes and long-range DNA interactions. Using an in vivo biotinylated nuclease-deficient Cas9 (dCas9) protein and programmable single guide RNAs (sgRNAs), this approach allows for high-resolution and locus-specific isolation of protein complexes and long-range chromatin looping associated with single copy CREs in mammalian cells. Unbiased analysis of the compositional structure of developmentally regulated or disease-associated CREs identifies new features of transcriptional regulation. Hence, CAPTURE provides a versatile platform to study genomic locus-regulating chromatin composition in a mammalian genome.
AB - Cis-regulatory elements (CREs) play a pivotal role in spatiotemporal control of tissue-specific gene expression, yet the molecular composition of the vast majority of CREs in native chromatin remains unknown. In this article, we describe the clustered regularly interspaced short palindromic repeats (CRISPR) affinity purification in situ of regulatory elements (CAPTURE) approach to simultaneously identify locus-specific chromatin-regulating protein complexes and long-range DNA interactions. Using an in vivo biotinylated nuclease-deficient Cas9 (dCas9) protein and programmable single guide RNAs (sgRNAs), this approach allows for high-resolution and locus-specific isolation of protein complexes and long-range chromatin looping associated with single copy CREs in mammalian cells. Unbiased analysis of the compositional structure of developmentally regulated or disease-associated CREs identifies new features of transcriptional regulation. Hence, CAPTURE provides a versatile platform to study genomic locus-regulating chromatin composition in a mammalian genome.
KW - chromatin
KW - cis-regulatory elements
KW - CRISPR
KW - DNA looping
KW - enhancer
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UR - http://www.scopus.com/inward/citedby.url?scp=85049582379&partnerID=8YFLogxK
U2 - 10.1002/cpmb.64
DO - 10.1002/cpmb.64
M3 - Article
C2 - 29927077
AN - SCOPUS:85049582379
VL - 123
JO - Current Protocols in Molecular Biology
JF - Current Protocols in Molecular Biology
SN - 1934-3639
IS - 1
M1 - e64
ER -