Leukemia pp 409-423

Part of the Methods in Molecular Biology™ book series (MIMB, volume 538)

Chromatin Immunoprecipitation (ChIP) for Analysis of Histone Modifications and Chromatin-Associated Proteins

Protocol

Summary

Disruption of epigenetic regulators of transcription is a central mechanism of oncogenesis. Many of the advances in the understanding of these mechanisms are attributable to the successful development of chromatin immunoprecipitation (ChIP) for in vivo detection of histone modifications as well as chromatin binding regulatory proteins. This is a powerful technique for analyzing histone modifications as well as binding sites for proteins that bind either directly or indirectly to DNA. Here we present two ChIP protocols. The first is particularly useful for identifying histone modifications or binding at specific, known genomic sites. The second, employing serial analysis of gene expression, is particularly powerful for the discovery of previously unidentified sites of modification or binding.

Key words:

Chromatin immunoprecipitation ChIP Histone modification Acetylation Methylation Quantitative PCR SAGE 

References

  1. 1.
    Orlando, V. (2000). Mapping chromosomal proteins in vivo by formaldehyde-crosslinked-chromatin immunoprecipitation. Trends Biochem Sci 25:99–104.PubMedCrossRefGoogle Scholar
  2. 2.
    Kirmizis, A. and Farnham, P.J. (2004). Genomic approaches that aid in the identification of transcription factor target genes. Exp Biol Med 229:705–721.Google Scholar
  3. 3.
    Huebert, D.J., Kamal, M., O’Donovan, A., and Bernstein, B.E. (2006). Genome-wide analysis of histone modifications by ChIP on chip. Methods 40:365–369.PubMedCrossRefGoogle Scholar
  4. 4.
    Shivaswamy, S. and Iyer, V.R. (2007). Genome analysis of chromatin status using tiling microarrays. Methods 41:304–311.PubMedCrossRefGoogle Scholar
  5. 5.
    Robyr, D., Kurdistani, S.K. and Grunstein, M. (2004). Analysis of genome-wide histone acetylation state and enzyme binding using DNA microarrays. Methods Enzymol 376:289–304.PubMedCrossRefGoogle Scholar
  6. 6.
    Saha, S., Sparks, A.B., Rago, C., Akmaev, V., Wang, C.J., Vogelstein, B., Kinzler, K.W. and Velculsecu, V.E. (2002). Using the transcriptome to annotate the genome. Nat Biotechnol 20:508–512.PubMedCrossRefGoogle Scholar
  7. 7.
    Roh, T.Y., Ngau, W.C., Cui, K., Landsman, D. and Zhao, K. (2004). High-resolution genome-wide mapping of histone modifications. Nat Biotechnol 22:1013–1016.PubMedCrossRefGoogle Scholar
  8. 8.
    Roh, T.Y., Cuddapah, S. and Zhao, K. (2005). Active chromatin domains are defined by acetylation islands revealed by genome-wide mapping. Genes Dev 19:542–552.PubMedCrossRefGoogle Scholar
  9. 9.
    Roh, T.Y., Cuddapah, S., Cui, K., and Zhao, K. (2006). The genomic landscape of histone modifications in human T cells. Proc Natl Acad Sci 103:15782–15787.PubMedCrossRefGoogle Scholar
  10. 10.
    Milne A. et al. (2005). Leukemogenic MLL fusion proteins bind across a broad region of the Hoxa9 locus, promoting transcription and multiple histone modifications. Cancer Res 65:11367–11374.Google Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  1. 1.Department of PathologyUniversity of Michigan Medical SchoolAnn ArborUSA

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