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J Vis Exp. 2017 Dec 12;(130). doi: 10.3791/56085.

Generation of Native Chromatin Immunoprecipitation Sequencing Libraries for Nucleosome Density Analysis.

Author information

1
Department of Microbiology and Immunology, Michael Smith Laboratories Centre for High-Throughput Biology, University of British Columbia.
2
Department of Microbiology and Immunology, Michael Smith Laboratories Centre for High-Throughput Biology, University of British Columbia; Canada's Michael Smith Genome Science Center, BC Cancer Agency; mhirst@bcgsc.ca.

Abstract

We present a modified native chromatin immunoprecipitation sequencing (ChIP-seq) experimental protocol compatible with a Gaussian mixture distribution based analysis methodology (nucleosome density ChIP-seq; ndChIP-seq) that enables the generation of combined measurements of micrococcal nuclease (MNase) accessibility with histone modification genome-wide. Nucleosome position and local density, and the posttranslational modification of their histone subunits, act in concert to regulate local transcription states. Combinatorial measurements of nucleosome accessibility with histone modification generated by ndChIP-seq allows for the simultaneous interrogation of these features. The ndChIP-seq methodology is applicable to small numbers of primary cells inaccessible to cross-linking based ChIP-seq protocols. Taken together, ndChIP-seq enables the measurement of histone modification in combination with local nucleosome density to obtain new insights into shared mechanisms that regulate RNA transcription within rare primary cell populations.

PMID:
29286469
PMCID:
PMC5755553
DOI:
10.3791/56085
[Indexed for MEDLINE]
Free PMC Article

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