An efficient chromatin immunoprecipitation (ChIP) protocol for studying histone modifications in Arabidopsis plants.
Chromatin immunoprecipitation (ChIP) is a powerful tool for the characterization of covalent histone modifications and DNA-histone interactions in vivo. The procedure includes DNA-histone cross-linking in chromatin, shearing DNA into smaller fragments, immunoprecipitation with antibodies against the histone modifications of interest, followed by PCR identification of associated DNA sequences. In this protocol, we describe a simplified and optimized version of ChIP assay by reducing the number of experimental steps and isolation solutions and shortening preparation times. We include a nuclear isolation step before chromatin shearing, which provides a good yield of high-quality DNA resulting in at least 15 mug of DNA from each immunoprecipitated sample (from 0.2 to 0.4 g of starting tissue material) sufficient to test > or =25 genes of interest. This simpler and cost-efficient protocol has been applied for histone-modification studies of various Arabidopsis thaliana tissues and is easy to adapt for other systems as well.
Saleh
Abdelaty
A
School of Biological Sciences, University of Nebraska, Lincoln, Nebraska 68588, USA. asaleh@ncsu.edu
Alvarez-Venegas
Raúl
R
Avramova
Zoya
Z
D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D018744:DNA, Plant / Q000235:genetics / Q000378:metabolism; D044127:Epigenesis, Genetic; D006657:Histones / Q000378:metabolism*
D029681:Arabidopsis Proteins; D018744:DNA, Plant; D006657:Histones