• 我要登录|
  • 免费注册
    |
  • 我的丁香通
    • 企业机构:
    • 成为企业机构
    • 个人用户:
    • 个人中心
  • 移动端
    移动端
丁香通 logo丁香实验_LOGO
搜实验

    大家都在搜

      大家都在搜

        0 人通过求购买到了急需的产品
        免费发布求购
        发布求购
        点赞
        收藏
        wx-share
        分享

        DamID: A Methylation-Based Chromatin Profiling Approach

        互联网

        1313
        Gene expression is a dynamic process and is tightly connected to changes in chromatin structure and nuclear organization (Schneider, R. and Grosschedl, R., 2007, Genes Dev. 21 , 3027–3043; Kosak, S. T. and Groudine, M., 2004, Genes Dev. 18 , 1371–1384). Our ability to understand the intimate interactions between proteins and the rapidly changing chromatin environment requires methods that will be able to provide accurate, sensitive, and unbiased mapping of these interactions in vivo (van Steensel, B., 2005, Nat. Genet. 37 Suppl , S18–24). One such tool is DamID chromatin profiling, a methylation-based tagging method used to identify the direct genomic loci bound by sequence-specific transcription factors, co-factors as well as chromatin- and nuclear-associated proteins genome wide (van Steensel, B. and Henikoff, S., 2000, Nat. Biotechnol . 18 , 424–428; van Steensel, Delrow, and Henikoff, 2001, Nat. Genet. 27 , 304–308). Combined with other functional genomic methods and bioinformatics analysis (such as expression profiles and 5C analysis), DamID emerges as a powerful tool for analysis of chromatin structure and function in eukaryotes. DamID allows the detection of the direct genomic targets of any given factor independent of antibodies and without the need for DNA cross-linking. It is highly valuable for mapping proteins that associate with the genome indirectly or loosely (e.g., co-factors). DamID is based on the ability to fuse a bacterial Dam-methylase to a protein of interest and subsequently mark the factor’s genomic binding site by adenine methylation. This marking is simple, highly specific, sensitive, inert, and can be done in both cell culture and living organisms. Below is a short description of the method, followed by a step-by-step protocol for performing DamID in Drosophila cells and embryos. Due to space limitations, the reader is referred to recent reviews that compare the method with other profiling techniques such as ChIP-chip as well as protocols for performing DamID in mammalian cells (NSouthall, T. D. and Brand, A. H., 2007, Nat. Struct. Mol. Biol. 14 , 869–871; Orian, A., 2006, Curr. Opin. Genet. Dev. 16 , 157–164; Vogel, M. J., Peric-Hupkes, D. and van Steensel, B. 2007, Nat. Protoc. 2 , 1467–1478).
        ad image
        提问
        扫一扫
        丁香实验小程序二维码
        实验小助手
        丁香实验公众号二维码
        扫码领资料
        反馈
        TOP
        打开小程序