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Related Experiment Videos

DNA methylation: bisulphite modification and analysis.

Susan J Clark1, Aaron Statham, Clare Stirzaker

  • 1Cancer Program, Garvan Institute of Medical Research, 384 Victoria St, Darlinghurst, Sydney, NSW 2010, Australia. s.clark@garvan.org.au

Nature Protocols
|April 5, 2007
PubMed
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This study presents a gold-standard bisulphite conversion protocol for analyzing DNA methylation. This method accurately quantifies DNA methylation at single-nucleotide resolution in mammalian genomes.

Area of Science:

  • Epigenetics
  • Molecular Biology
  • Genomics

Background:

  • DNA methylation is a crucial epigenetic modification in mammalian genomes.
  • Methylation patterns are vital for development and tissue differentiation.
  • Aberrant DNA methylation is linked to various diseases, including cancer.

Purpose of the Study:

  • To introduce a gold-standard bisulphite conversion protocol.
  • To enable accurate and reproducible analysis of DNA methylation sequences.
  • To determine and quantify DNA methylation at single-nucleotide resolution.

Main Methods:

  • Utilizes bisulphite treatment to differentiate cytosine and 5-methylcytosine (5-MeC).
  • Exploits differential deamination sensitivity under acidic conditions.

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  • Protocol adaptable for single tubes or 96-well formats.

    • Main Results:

    • The bisulphite conversion protocol allows for single-nucleotide resolution of DNA methylation.
    • Accurate quantification of methylation states in genes or genomic regions is achievable.
    • The protocol can be completed within 8 to 18 hours.

    Conclusions:

    • The described bisulphite conversion protocol is a reliable method for DNA methylation analysis.
    • This technique is essential for understanding the biological functions of methylation changes.
    • It provides a foundation for further research in epigenetics and disease states.