Isolation of highly purified yeast nuclei for nuclease mapping of chromatin structure

Joseph C. Reese, Hesheng Zhang, Zhengjian Zhang

Research output: Chapter in Book/Report/Conference proceedingChapter

2 Scopus citations

Abstract

Probing chromatin structure with nucleases is a well-established method for determining the accessibility of DNA to gene regulatory proteins and measuring competency for transcription. A hallmark of many silent genes is the presence of translationally positioned nucleosomes over their promoter regions, which can be inferred by the sensitivity of the underlying DNA to nucleases, particularly micrococcal nuclease. The quality of this data is highly dependent upon the nuclear preparation, especially if the digestion products are analyzed by high-resolution detection methods such as reiterative primer extension. Here we describe a method to isolate highly purified nuclei from the budding yeast Saccharomyces cerevisiae and the use of micrococcal nuclease to map the positions of nucleosomes at the RNR3 gene. Nuclei isolated by this procedure are competent for many of the commonly used chromatin mapping and detection procedures.

Original languageEnglish (US)
Title of host publicationThe Nucleus
Subtitle of host publicationVolume 1: Nuclei and Subnuclear Components
Pages43-53
Number of pages11
DOIs
StatePublished - 2008

Publication series

NameMethods in Molecular Biology
Volume463
ISSN (Print)1064-3745

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Genetics

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    Reese, J. C., Zhang, H., & Zhang, Z. (2008). Isolation of highly purified yeast nuclei for nuclease mapping of chromatin structure. In The Nucleus: Volume 1: Nuclei and Subnuclear Components (pp. 43-53). (Methods in Molecular Biology; Vol. 463). https://doi.org/10.1007/978-1-59745-406-3-3