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Purification of immune-active macrophage super enhancers by chemical cross-linked chromatin immune precipitation.

ABSTRACT: Isolation of extraordinarily long-length super-enhancers (SEs) using typical chromatin immune precipitation (ChIP) techniques can lead to DNA breakage due to uncontrolled cross-linking. We present a redefined ChIP technique for SE purification. After controlled paraformaldehyde-based cross-linking, glycine was used to quench the cross-linker followed by mild sonication. The sonication produced ideal fragment length of long-length SE chromatin. Presently, miR146a-5p SE of macrophages was pulled using BRD4 protein. Our protocol can reproducibly simplify the SE element isolation issues, in a quality-controlled manner. For complete details on the use and execution of this protocol, please refer to Das et al. (2021).¹.

SUBMITTER: Das S

PROVIDER: S-EPMC9852664 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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Purification of immune-active macrophage super enhancers by chemical cross-linked chromatin immune precipitation.

Das Sonali S Mukherjee Sohitri S Ali Nahid N

STAR protocols 20230111 1

Isolation of extraordinarily long-length super-enhancers (SEs) using typical chromatin immune precipitation (ChIP) techniques can lead to DNA breakage due to uncontrolled cross-linking. We present a redefined ChIP technique for SE purification. After controlled paraformaldehyde-based cross-linking, glycine was used to quench the cross-linker followed by mild sonication. The sonication produced ideal fragment length of long-length SE chromatin. Presently, miR146a-5p SE of macrophages was pulled u ...[more]

PMID: 36638018

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