Immunofluorescence Imaging Strategy for Evaluation of the Accessibility of DNA 5-Hydroxymethylcytosine in Chromatins

CORC > 生态环境研究中心 > 中国科学院生态环境研究中心 > 环境化学与生态毒理学国家重点实验室

	Immunofluorescence Imaging Strategy for Evaluation of the Accessibility of DNA 5-Hydroxymethylcytosine in Chromatins
	Wang, Hailin; Zhong, Shangwei; Li, Zhe; Jiang, Ting; Li, Xiangjun
刊名	ANALYTICAL CHEMISTRY
	2017-11-07
卷号	89 期号:11 页码:5703-5707
英文摘要	DNA 5-hydroxymethylcytosine (5hmC) is an important epigenetic modification found in various mammalian cells. Immunofluorescence imaging analysis essentially provides visual pictures for the abundance and distribution of DNA 5hmC in single cells. However, nuclear DNA is usually wrapped around nucleosomes, packaged into chromatins, and further bound with many functional proteins. These physiologically relevant events would generate barriers to the anti-5hmC antibody to selectively recognize 5hmC in DNA. By taking advantage of these naturally generated barriers, here, we present a strategy to evaluate the accessibility of DNA 5hmC in chromatins in situ. We demonstrate that a few of the 5hmC sites in DNA are exposed or accessible to anti-5hmC antibody under nondenaturing conditions, suggesting that these 5hmC sites are not covered by functional DNA-binding proteins in mouse embryonic stem cells. Consistently, these 5hmC foci were distributed in open euchromatin regions as revealed by the 4',6-diamidino-2-phenylindole (DAPI) staining. By overexpressing TET1 catalytic domain (responsible for oxidation 5mC to produce 5hmC) in human MCF-7 cells, we observed a significant increase in accessible 5hmC along with an increase in total 5hmC sites. Collectively, by the use of the nondenaturing immunofluorescence imaging approach, we could obtain a visual landscape on the accessibility of DNA 5hmC in chromatins.
内容类型	期刊论文
源URL	[http://ir.rcees.ac.cn/handle/311016/39337]
专题	生态环境研究中心_环境化学与生态毒理学国家重点实验室
推荐引用方式 GB/T 7714	Wang, Hailin,Zhong, Shangwei,Li, Zhe,et al. Immunofluorescence Imaging Strategy for Evaluation of the Accessibility of DNA 5-Hydroxymethylcytosine in Chromatins[J]. ANALYTICAL CHEMISTRY,2017,89(11):5703-5707.
APA	Wang, Hailin,Zhong, Shangwei,Li, Zhe,Jiang, Ting,&Li, Xiangjun.(2017).Immunofluorescence Imaging Strategy for Evaluation of the Accessibility of DNA 5-Hydroxymethylcytosine in Chromatins.ANALYTICAL CHEMISTRY,89(11),5703-5707.
MLA	Wang, Hailin,et al."Immunofluorescence Imaging Strategy for Evaluation of the Accessibility of DNA 5-Hydroxymethylcytosine in Chromatins".ANALYTICAL CHEMISTRY 89.11(2017):5703-5707.