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Bisulfite-Based Methylation Analysis of Imprinted Genes

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Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 181))

Abstract

Genomic imprinting is an epigenetically controlled form of gene regulation leading to the preferential expression of one parental gene copy. To date, approximately 40 imprinted genes have been described that are exclusively or predominantly expressed from either the paternal or the maternal allele (www.mgu.har.mrc.ac.uk/imprinting/implink.html). Changes in the imprinted expression of such genes result in developmental abnormalities; in the human they are associated with several diseases and various types of cancer(1-3).

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References

  1. Constancia, M., Pickard, B., Kelsey, G., and Reik, W. (1998) Imprinting mechanisms. Genome Res. 8, 881–900

    PubMed  CAS  Google Scholar 

  2. Nicholls, R. D., Saitoh, S., and Horsthemke, B. (1998) Imprinting in Prader-Willi and Angelman syndromes. TIGS 14, 194–200.

    Article  CAS  Google Scholar 

  3. Tycko, B. (1997) DNA methylation in genomic imprinting. Mutat. Res.-Rev. in Mutat. Res. 386, 131–140.

    CAS  Google Scholar 

  4. Reik, W.and Walter, J. (2001) Genomic imprinting; parental influence on the genome. Nature Rev. Genet. 2, 21–32.

    Article  PubMed  CAS  Google Scholar 

  5. Li, E. B. C.and Jaenisch, R. (1992) Targeted mutation of the DNA methyltransferase gene results in embryonic lethality. Cell 69, 915–926.

    Article  PubMed  CAS  Google Scholar 

  6. Frommer, M., McDonald, L. E., Millar, D. S., Collis, C. M., Watt, F., Grigg, G.W., Molloy, P. L., and Paul, C. L. (1992) A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands. Proc. Natl. Acad. Sci.USA 89, 1827–1831.

    Article  PubMed  CAS  Google Scholar 

  7. Paul, C. L.and Clark, S. J. (1996) Cytosine methylation: Quantitation by automated genomic sequencing and GENESCAN analysis. BioTechniques 21, 126–133.

    PubMed  CAS  Google Scholar 

  8. Gonzalgo, M.and Jones, P. A. (1997) Rapid quantitation of methylation differences at specific sites using methylation-sensitive single nucleotide primer extension (Ms-SNuPE) Nucleic Acids Res. 25, 2529–2531.

    Article  PubMed  CAS  Google Scholar 

  9. Clark, S. J., Harrison, J., Paul, C. L., and Frommer, M. (1994) High sensitivity mapping of methylated cytosines. Nucl. Acids Res. 22, 2990–2997

    Article  PubMed  CAS  Google Scholar 

  10. Feil, R., Walter, J., Allen, 6N. D., and Kelsey, G. (1994) Developmental control of allelic methylation in the imprinted mouse Igf2 and H19 genes. Development 120, 2933–2943

    PubMed  CAS  Google Scholar 

  11. Raizis, A. M., Schmitt, F., and Jost, J. P. (1994) A bisulfite method of 5-methylcytosine mapping that minimises template degradation. Anal. Biochem. 226, 161–166.

    Article  Google Scholar 

  12. Olek, A. Oswald, J., and Walter, J. (1996) A modified and improved method for bisulfite based cytosine methylation analysis. Nucleic Acids Res. 24, 5064–5066.

    Article  PubMed  CAS  Google Scholar 

  13. Paulin, R., Grigg, G.W., Davey, M. W., and Piper, A. A. (1998) Urea improves efficiency of bisulfite-mediated sequencing of 5-methylcytosine in genomic DNA. Nucl. Acids Res. 26, 5009–5010.

    Article  PubMed  CAS  Google Scholar 

  14. Hayatsu, H., Wataya, Y., Kai, K., and Iida, S. (1970) Reaction of sodium bisulfite with uracil, cytosine, and their derivatives. Biochemistry 9, 2858–2864.

    Article  PubMed  CAS  Google Scholar 

  15. Wang, R. Y.-H., Gehrke, C. W., and Ehrlich, M. (1980) Comparison of bisulfite modification of 5-methyldeoxycytidine and deoxycytidine residues. Nucleic Acids Res. 8, 4777–4790.

    Article  PubMed  CAS  Google Scholar 

  16. Sambrook, G., Fritsch, E. F., and Maniatis, T. (1988) Molecular Cloning and Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.

    Google Scholar 

  17. Warnecke, P. M., Stirzaker, C., Melki, J. R., Millar, D. S., Paul, C. L., and Clark, S. J.(1997) Detection and measurement of PCR bias in quantitative methylation analysis of bisulfite-treated DNA. Nucl. Acids Res. 25, 4422–4426.

    Article  PubMed  CAS  Google Scholar 

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© 2002 Humana Press Inc., Totowa, NJ

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Engemann, S., El-Maarri, O., Hajkova, P., Oswald, J., Walter, J. (2002). Bisulfite-Based Methylation Analysis of Imprinted Genes. In: Ward, A. (eds) Genomic Imprinting. Methods in Molecular Biology™, vol 181. Humana Press. https://doi.org/10.1385/1-59259-211-2:217

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  • DOI: https://doi.org/10.1385/1-59259-211-2:217

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-741-0

  • Online ISBN: 978-1-59259-211-1

  • eBook Packages: Springer Protocols

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