Skip to main content
Journal of Clinical Laboratory Analysis logoLink to Journal of Clinical Laboratory Analysis
. 2009 May 19;23(3):172–174. doi: 10.1002/jcla.20314

Simplified and versatile method for bisulfite‐based DNA methylation analysis of small amounts of DNA

SK Tiwari 1, G Manoj 1, K Prasanth 1, G Sivaram 1, VK Sharma 1, MA Habeeb 1, AA Khan 1,, CM Habibullah 1
PMCID: PMC6648936  PMID: 19455637

Abstract

Epigenetic alterations of gene function play a central role in the pathogenesis of many tumors and in the process of aging. Abnormal methylation at transcriptional sites of genes results in epigenetic silencing of the genes that protect against tumor formation or that repair DNA. To date, several studies have analyzed methylation status by oligonucleotide arrays, restriction analysis (COBRA), methylation‐specific amplification, and sequence analysis. Thisrequires high concentration of bisulfite‐treated DNA, which mandates use of commercially available expensive kits, and is an often laborious and time‐consuming task. In this article, we report a simplified high‐throughput method, which can serve as a surrogate for screening methylation profiles of various genes and has high sensitivity compared with the other methods described previously. J. Clin. Lab. Anal. 23:172–174, 2009. © 2009 Wiley‐Liss, Inc.

Keywords: bisulfite modification, DNA methylation, epigenetic changes, gene silencing

REFERENCES

  • 1. Jones PA, Takai D. The role of DNA methylation in mammalian epigenetics. Science 2001;293:1068–1070. [DOI] [PubMed] [Google Scholar]
  • 2. Delgado S, Gomez M, Bird A, et al. Initiation of DNA replication at CpG islands in mammalian chromosomes. EMBO J 1998;17:2426–2435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3. Jones PA, Laird PW. Cancer epigenetic comes of age. Nat Genet 1999;21:163–167. [DOI] [PubMed] [Google Scholar]
  • 4. Kang GH, Shim Y‐H, Jung W‐Y, et al. CpG island methylation in premalignant stages of gastric carcinoma. Cancer Res 2001;61:2285–2847. [PubMed] [Google Scholar]
  • 5. Frommer M, McDonald LE, Millar DS, et al. A genomic sequencing protocol that yields a positive display of 5‐methylcytosine residues in individual DNA strands. Proc Natl Acad Sci USA 1992;89:1827–1831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6. Raizis AM, Schmitt F, Jost JP. A bisulfite method of 5‐methylcytosine mapping that minimizes template degradation. Anal Biochem 1995;226:161–166. [DOI] [PubMed] [Google Scholar]
  • 7. Tyler LN, Ai L, Zuo C, et al. Analysis of promoter hypermethylation of death‐associated protein kinase and p16 tumor suppressor genes in actinic keratoses and squamous cell carcinomas of the skin. Mod Pathol 2003;16:660–664. [DOI] [PubMed] [Google Scholar]
  • 8. Olek A, Oswald J, Walter J. A modified and improved method for bisulphite based cytosine methylation analysis. Nucleic Acids Res 1996;24:5064–5066. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9. Sambrook J, Russell DW. Molecular Cloning. A Laboratory Manual, third edition, Vol. 1 Cold Spring Harbor, NY: Cold Spring Harbor Laboratory, 2001. p 6.61–6.62. [Google Scholar]
  • 10. Cone RW, Huang ML, Ashley R, et al. Human herpesvirus 6 DNA in peripheral blood cells and saliva from immunocompetent individual. J Clin Microbiol 1993;31:1262–1267. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Clinical Laboratory Analysis are provided here courtesy of Wiley

RESOURCES