Cited by 358Epigenetics and DNA MethylationGenetic Syndromes and ImprintingDNA and Nucleic Acid Chemistry

DNA Methylation Detection: Bisulfite Genomic Sequencing Analysis

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Li, Yuanyuan, Tollefsbol, Trygve O.·2011·Methods in Molecular Biology

Abstract

DNA methylation, which most commonly occurs at the C5 position of cytosines within CpG dinucleotides, plays a pivotal role in many biological procedures such as gene expression, embryonic development, cellular proliferation, differentiation, and chromosome stability. Aberrant DNA methylation is often associated with loss of DNA homeostasis and genomic instability leading to the development of human diseases such as cancer. The importance of DNA methylation creates an urgent demand for effective methods with high sensitivity and reliability to explore innovative diagnostic and therapeutic strategies. Bisulfite genomic sequencing developed by Frommer and colleagues was recognized as a revolution in DNA methylation analysis based on conversion of genomic DNA by using sodium bisulfite. Besides various merits of the bisulfite genomic sequencing method such as being highly qualitative and quantitative, it serves as a fundamental principle to many derived methods to better interpret the mystery of DNA methylation. Here, we present a protocol currently frequently used in our laboratory that has proven to yield optimal outcomes. We also discuss the potential technical problems and troubleshooting notes for a variety of applications in this field.

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  • Bisulfite sequencing is used to identify methylated DNA by converting all unmethylated cytosine bases to uracil through the addition of a bisulfite agent such as sodium bisulfite.

    “Bisulfite sequencing is used to identify methylated DNA (5mC) by converting all unmethylated cytosine bases to uracil through the addition of a bisulfate agent such as sodium bisulfate.”

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  1. [21]Paulin R Grigg GW Davey MW Piper AA 1998 Urea improves efficiency of bisulphite-mediated sequencing of 59-methylcytosine in genomic DNA Nucleic Acids Res 26 5009 5010 9776768 PMC147923

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Authors
Li, Y., & Tollefsbol, T. O.
Affiliations
University of Alabama at Birmingham (US)
Corresponding author
Tollefsbol, Trygve O.
Venue
Methods in Molecular Biology
Publisher
Humana Press
Published
2011
Volume
791
Pages
11-21
ISSN
  • 1064-3745 (ISSN-L)
  • 1940-6029
OpenAlex
W1522996100
Cited by
352
References
26
Citations by year
2026: 25 · 2025: 34 · 2024: 41 · 2023: 32 · 2022: 35 · 2021: 41 · 2020: 36 · 2019: 23 · 2018: 23 · 2017: 22 · 2016: 16 · 2015: 13 · 2014: 6 · 2013: 3 · 2012: 1
FWCI
4.57
Type
book chapter
Language
en
Open access
Locations
  • Methods in molecular biology (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
Funders
  • Susan G. Komen
  • National Cancer Institute (R01CA129415)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Epigenetics and DNA Methylation
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genetic Syndromes and Imprinting
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
Keywords
DNA methylation, bisulfite sequencing, CpG, genomic DNA, aberrant methylation, genomic instability, cancer, troubleshooting
MeSH terms
Base Sequence, Cloning, Molecular, DNA, Laboratories, Sulfites, Polymerase Chain Reaction, Sequence Analysis, DNA, DNA Methylation, Genomics

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