Abstract
The methylated DNA immunoprecipitation microarray (MeDIP-chip) is a genome-wide, high-resolution approach to detect DNA methylation in whole genome or CpG (cytosine base followed by a guanine base) islands. The method utilizes anti-methylcytosine antibody to immunoprecipitate DNA that contains highly methylated CpG sites. Enriched methylated DNA can be interrogated using DNA microarrays or by massive parallel sequencing techniques. This combined approach allows researchers to rapidly identify methylated regions in a genome-wide manner, and compare DNA methylation patterns between two samples with diversely different DNA methylation status. MeDIP-chip has been applied successfully for analyses of methylated DNA in the different targets including animal and plant tissues (1, 2). Here we present a MeDIP-chip protocol that is routinely used in our laboratory, illustrated with specific examples from MeDIP-chip analysis of breast cancer cell lines. Potential technical pitfalls and solutions are also provided to serve as workflow guidelines.
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References
Zilberman, D., Gehring, M., Tran, R.K., Ballinger, T., Henikoff, S. (2007) Genome-wide analysis of Arabidopsis thaliana DNA methylation uncovers an interdependence between methylation and transcription. Nat. Genet. 39, 61–69.
Rauch, T., Wang, Z., Zhang, X., Zhong, X., Wu, X., Lau, S. K., Kernstine, K. H., Riggs, A. D., Pfeifer, G. P., (2007) Homeobox gene methylation in lung cancer studied by genome-wide analysis with a microarray-based methylated CpG island recovery assay. Proc. Natl. Acad. Sci. 104, 5527–5532.
Esteller, M. (2008) Epigenetics in Cancer. N. Engl. J. Med. 358, 1148–1159.
Liu, K., Wang, Y. F., Cantemir, C., Muller, M. T. (2003) Endogenous assays of DNA methyltransferases: Evidence for differential activities of DNMT1, DNMT2, and DNMT3 in mammalian cells in vivo. Mol. Cell Biol. 23, 2709–19.
Rollings, R.A., Haghighi, F., Edwards, J.R., Das, R., Zhang, M.Q., Ju, J., Bestor, T.H. (2006) Large-scale structure of genomic methylation patterns. Genome Res. 16, 157–163.
Laird, P.W. (2005) Cancer epigenetics. Hum. Mol. Genet. 14, R65–76.
Bird, A. (2002) DNA methylation patterns and epigenetic memory. Genes Dev. 16, 6–21.
Jaenisch, R., Bird, A. (2003) Epigenetic regulation of gene expression: how the genome integrates intrinsic and environmental signals. Nat. Genet. 33, 245–254.
Das, P. M., Signal, R. (2004) DNA methylation and cancer. J. Clin. Oncol. 22, 4632–4642.
Feinberg, A. P., Tycko, B. (2004) The history of cancer epigenetics. Nat. Rev. Cancer 4, 143–153.
Suzuki, M. M., Bird, A. (2008) DNA methylation landscapes: provocative insights from epigenomics. Nat. Rev. Genet. 9, 465–476.
Herman, J. G., Graff, J. R., Myohanen, S. et al. (1996) Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. Proc. Natl. Acad. Sci. USA 93, 9821–9826.
Eads, C. A., Danenberg, K. D., Kawakami, K., et al. (1999) CpG island hypermethylation in human colorectal tumors is not associated with DNA methyltransferase overexpression. Cancer Res. 59, 2302–2306.
Xion, Z., Laird, P. W. (1997) COBRA: a sensitive and quantitative DNA methylation assay. Nucleic Acids Res. 25, 2532–2534.
Gonzalgo, M. L., Jones, P. A. (1997) Rapid quantitation of methylation differences at specific sites using methylation-sensitive single nucleotide primer extension (Ms-SNuPE). 25, 2529–2531.
Gitan, R. S., Shi, H., Chen, C. M., Yan, P. S., Huang, T. H. (2002) Methylation-specific oligonucleotides microarray: a new potential for high-throughout methylation analysis. Genome Res. 12, 158–164.
Yan, P. S., Chen, C. M., Shi, H., Rahmatpanah F., Wei, S. H., Huang, T. H. (2002) Applications of CpG island microarrays for high-throughout analysis of DNA methylation. J. Nutr. 132, 2430S–2434S.
Weber, M., Davies, J. J., Wittig, D., Oakeley, E. L., Haase, M., Lam, W. L., Schübeler, D. (2005) Chromosome-wide and promoter-specific analyses identify sites of differential DNA methylation in normal and transformed human cells. Nat. Genet. 37, 853–862.
Lee, T.I., Johnstone, S.E., Young, R.A., (2006) Chromatin immunoprecipitation and microarray-based analysis of protein location. Nature Protocols 1, 729–748.
Down, T.A., Rakyan, V.K., Turner, D.J., Flicek, P., Li, H., Kulesha, E., Graf, S., Johnson, N., Herrero, J., Tomazou, E.M., Thorne, N.P., Backdahl, L., Herberth, M., Howe, K.L., Jackson, D.K., Miretti, M.M., Marioni, J.C., Birney, E., Hubbard, T.J.P., Durbin, R., Tavare, S., Beck, S., (2008) A Bayesian deconvolution strategy for immunoprecipitation-based DNA methylome analysis. Nat. Biotech. 26, 779–785.
Jacinto, F.V., Ballestar, E., Esteller, M., (2008) Methyl-DNA immunoprecipitation (MeDIP): hunting down the DNA methylation. Biotechniques 44, 35–43.
Kininis, M., Chen, B.S., Diehl, A.G., Isaacs, G.D., Zhang, T., Siepel, A.C., Clark, A.G., Kraus, W.L., (2007) Genomic analyses of transcription factor binding, histone acetylation, and gene expression reveal mechanistically distinct classes of estrogen-regulated promoters. Mol. Cell. Biol. 27, 5090–5104.
Liu, C.L., Schreiber, S.L., Bernstein, B.E. (2003) Development and validation of a T7 based linear amplification for genomic DNA. BMC Genomics 4, 19.
Schilling, E., Rehli, M. (2007) Global, comparative analysis of tissue-specific promoter CpG methylation. Genomics. 90, 314–323.
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Weng, YI., Huang, T.HM., Yan, P.S. (2009). Methylated DNA Immunoprecipitation and Microarray-Based Analysis: Detection of DNA Methylation in Breast Cancer Cell Lines. In: Park-Sarge, OK., Curry, T. (eds) Molecular Endocrinology. Methods in Molecular Biology, vol 590. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60327-378-7_10
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DOI: https://doi.org/10.1007/978-1-60327-378-7_10
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