Abstract
The characterization of protein-deoxyribonucleic acid (DNA) interactions occurring at an allele-specific level is important to resolving the functional consequences of genetic variation in non-coding DNA for gene expression and regulation. The approach of haplotype-specific chromatin immunoprecipitation (i.e., haploChIP) resolves in living cells relative protein-DNA binding to a particular allele through immunoprecipitation of proteins crosslinked to DNA. Single-nucleotide polymorphisms present in a heterozygous form are used as markers to differentiate allelic origin. This in turn allows resolution of specific haplotypes showing differences in relative protein occupancy. The haploChIP approach allows testing of in vitro hypotheses that a transcription factor protein shows haplotype specific occupancy. In addition, the haploChIP approach allows screening of haplotypes for differences in relative gene expression by immunoprecipitation using antibodies to phosphorylated Pol II.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Knight, J. C., Keating, B. J., Rockett, K. A., and Kwiatkowski, D. P. (2003) In vivo characterization of regulatory polymorphisms by allele-specific quantification of RNA polymerase loading. Nat. Genet. 33, 469–475.
Glazier, A. M., Nadeau, J. H., and Aitman, T. J. (2002) Finding genes that underlie complex traits. Science 298, 2345–2349.
Cargill, M., Altshuler, D., Ireland, J., et al. (1999) Characterization of single-nucleotide polymorphisms in coding regions of human genes. Nat. Genet. 22, 231–238.
Knight, J. C. (2003) Functional implications of genetic variation in non-coding DNA for disease susceptibility and gene regulation. Clin. Sci. (Lond) 104, 493–501.
Yan, H., Yuan, W., Velculescu, V. E., Vogelstein, B., and Kinzler, K. W. (2002) Allelic variation in human gene expression. Science 297, 1143.
Knight, J. C., Keating, B. J., and Kwiatkowski, D. P. (2004) Allele-specific repression of lymphotoxin-alpha by activated B cell factor-1. Nat. Genet. 36, 394–399.
Orlando, V., Strutt, H., and Paro, R. (1997) Analysis of chromatin structure by in vivo formaldehyde cross-linking. Methods 11, 205–214.
Jurinke, C., van den Boom, D., Cantor, C. R., and Koster, H. (2002) Automated genotyping using the DNA MassArray technology. Methods Mol. Biol. 187, 179–192.
Braun, A., Little, D. P., and Koster, H. (1997) Detecting CFTR gene mutations by using primer oligo base extension and mass spectrometry. Clin. Chem. 43, 1151–1158.
Takahashi, Y., Rayman, J. B., and Dynlacht, B. D. (2000) Analysis of promoter binding by the E2F and pRB families in vivo: distinct E2F proteins mediate activation and repression. Genes Dev. 14, 804–816.
Cowles, C. R., Joel, N. H., Altshuler, D., and Lander, E. S. (2002) Detection of regulatory variation in mouse genes. Nat. Genet. 32, 432–437.
Weber, M., Hagege, H., Lutfalla, G., et al. (2003) A real-time polymerase chain reaction assay for quantification of allele ratios and correction of amplification bias. Anal. Biochem. 320, 252–258
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2005 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Knight, J.C. (2005). HaploChIP. In: Innocenti, F. (eds) Pharmacogenomics. Methods in Molecular Biology™, vol 311. Humana Press. https://doi.org/10.1385/1-59259-957-5:049
Download citation
DOI: https://doi.org/10.1385/1-59259-957-5:049
Publisher Name: Humana Press
Print ISBN: 978-1-58829-440-1
Online ISBN: 978-1-59259-957-8
eBook Packages: Springer Protocols