Abstract
Development of multicellular organisms is based on specialized gene expression programs. Because chromatin establishes the environment for transcription, understanding composition and dynamics of chromatin is an important part of developmental biology. The knowledge about chromatin has been greatly advanced by the chromatin immunoprecipitation (ChIP) technique, because ChIP allows to map the position of proteins as well as modifications of DNA and histones to specific genomic regions. Although ChIP has been applied to a wide range of model organisms, including Arabidopsis, it remains a challenging technique, and a careful experimental setup including appropriate positive and negative controls are required to obtain reliable results. Here, we describe a ChIP protocol adapted for material from Arabidopsis, which we routinely apply in our laboratory, and we discuss required controls and methods for data analysis.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsSpringer Nature is developing a new tool to find and evaluate Protocols. Learn more
References
Luger, K., Mader, A. W., Richmond, R. K., Sargent, D. F., and Richmond, T. J. (1997) Crystal structure of the nucleosome core particle at 2.8 A resolution. Nature 389, 251–260.
Grunstein, M. (1997) Histone acetylation in chromatin structure and transcription. Nature 389, 349–352.
Duo, Y., Mizzen, C. A., Abrams, M., Allis, C. D., and Gorovsky, M. A. (1999) Phosphorylation of linker histone H1 regulates gene expression in vivo by mimicking H1 removal. Mol Cell 4, 641–647.
Zhang, Y. and Reinberg, D. (2001) Transcription regulation by histone methylation: interplay between different covalent modifications of the core histone tails. Genes Dev 15, 2343–2360.
Gehring, M. and Henikoff, S. (2007) DNA methylation dynamics in plant genomes. Biochem Biophys Acta 1769, 276–286.
Gilmour, D. S. and Lis, T. J. (1984) Detecting protein-DNA interactions in vivo: Distribution of RNA polymerase on specific bacterial genes. Proc Natl Acad Sci USA 81, 4275–4279.
Dedon, P. C., Soults, J. A., Allis, C. D., and Gorovsky, M. A. (1991) A simplified formaldehyde fixation and immunoprecipitation technique for studying protein–DNA interactions. Anal Biochem 197, 83–90.
Gendrel, A.-V., Lippman, Z., Yordan, C., Colot, V., and Martienssen, R. A. (2002) Dependence of heterochromatic histone H3 methylation patterns on the Arabidopsis gene DDM1. Science 297, 1871–1873.
Gendrel, A.-V., Lippman, Z., Martienssen, R. A., and Colot, V. (2005) Profiling histone modification patterns in plants using genomic tiling microarrays. Nat Methods 2, 213–218.
Makarevich, G., Leroy, O., Akinci, U., Schubert, D., Clarenz, O., Goodrich, J., Grossniklaus, U., and Köhler, C. (2006) Different Polycomb group complexes regulate common target genes in Arabidopsis. EMBO Rep 7, 947–952.
Schönrock, N., Bouveret, R., Leroy, O., Borghi, L., Köhler, C., Gruissem, W., and Hennig, L. (2006) Polycomb-group proteins repress the floral activator AGL19 in the FLC-independent vernalization pathway. Genes Dev 20, 1667–1678.
Villar, C. B. R., Erilova, A., Makarevich, G., Trösch, R., and Köhler, C. (2009) Control of PHERES1 imprinting in Arabidopsis by direct tandem repeats. Mol Plant 2, 654–660.
O’Neill, L. P. and Turner, B. M. (2003) Immunoprecipitation of native chromatin: NChIP. Methods 31, 76–82.
Kurdistani, S. K. and Grunstein, M. (2003) In vivo protein-protein and protein-DNA crosslinking for genome-wide binding microarray. Methods 31, 90–95.
Fujita, N., Jaye, D. L., Kajita, M., Geigerman, C., Moreno, C. S., and Wade, P. A. (2003) MTA3, a Mi-2/NuRD complex subunit regulates an invasive growth pathway in breast cancer. Cell 113, 207–219.
Nowak, D. E., Tian, B., and Brasier, A. R. (2005) Two-step crosslinking method for the identification of NF-kappa B gene network by chromatin immunoprecipitation. BioTechniques 39, 715–725.
Chua, Y. L., Mott, E., Brown, A. P. C., MacLean, D., and Gray, J. C. (2004) Microarray analysis of chromatin-immunoprecipitated DNA identifies specific regions of tobacco genes associated with acetylated histones. Plant J 37, 789–900.
Zhang, X., Clarenz, O., Cokus, S., Bernatavichute, Y. V., Pellegrini, M., Goodrich, J., and Jacobsen, S. E. (2007) Whole-genome analysis of histone H3 lysine 27 trimethylation in Arabidopsis. PLoS Biol 5, e129.
Jothi, R., Cuddapah, S., Barski, A., Cui, K., and Zhao, K. (2008) Genome-wide identification of in vivo protein–DNA binding sites from ChIP-seq data.Nucleic Acids Res 36, 5221–5231.
Haring, M., Offermann, S., Danker, T., Horst, I., Peterhänsel, C., and Stam, M. (2007) Chromatin immunoprecipitation: Optimization, quantitative analysis and data normalization. Plant Methods 3, 11.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2010 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Villar, C.B., Köhler, C. (2010). Plant Chromatin Immunoprecipitation. In: Hennig, L., Köhler, C. (eds) Plant Developmental Biology. Methods in Molecular Biology, vol 655. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-765-5_27
Download citation
DOI: https://doi.org/10.1007/978-1-60761-765-5_27
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-60761-764-8
Online ISBN: 978-1-60761-765-5
eBook Packages: Springer Protocols