Summary: DNA methylation is an epigenetic event that affects cell function by altering gene expression and refers to the covalent addition of a methyl group, catalyzed by DNA methyltransferase (DNMT), to the 5-carbon of cytosine in a CpG dinucleotide. Methods for DNA methylation analysis can be divided roughly into two types: global and gene-specific methylation analysis. For global methylation analysis, there are methods which measure the overall level of methyl cytosines in genome such as chromatographic methods and methyl accepting capacity assay. For gene-specific methylation analysis, a large number of techniques have been developed. Most early studies used methylation sensitive restriction enzymes to digest DNA followed by Southern detection or PCR amplification. Recently, bisulfite reaction based methods have become very popular such as methylation specific PCR (MSP), bisulfite genomic sequencing PCR. Additionally, in order to identify unknown methylation hot-spots or methylated CpG islands in the genome, several of genome-wide screen methods have been invented such as Restriction Landmark Genomic Scanning for Methylation (RLGS-M), and CpG island microarray.
DNA Methylation Analysis(Methods-Online) Very nice protocol on DNA methylation mapping with backgroud information about the procedures. Three methods are described: 1)Methylation analysis using restriction enzyme digestion; 2)Bisulphite (or bisulfite)genomic sequencing; 3)Single nucleotide primer extension (SNuPE).
http://www.methods-online.net/methods/DNAmethylati... Added: Wed Sep 04 2002, Hits: 6938, Reviews: 0Write reviewCached
(Issa Lab) This assay can be used to measure methyltransferase activity in small, snap frozen pieces of tissue (e.g. colon polyps), or in cultured cells http://www.mdanderson.org/departments/methylation/... Added: Tue May 14 2002, Hits: 1940, Reviews: 0Write reviewCached
Methyltransferase-based single-promoter analysis assay (MSPA)
(Epigenome Network of Excellence) Methylation-sensitive promoter analysis allows for the study of unmethylated CpG islands by treatment of nuclei with the CpG-specific DNA methyltransferase SssI (M.SssI), followed by genomic bisulfite sequencing of individual progeny DNA molecules. This gives single molecule resolution over the promoter and allows for the physical linkage between binding sites on individual promoter molecules to be maintained. http://www.epigenome-noe.net/researchtools/protoco... Added: Sat Jan 31 2009, Hits: 508, Reviews: 0Write reviewCached
(Makoto Kawase) Detailed Restriction Landmark Genomic Scanning (RLGS) protocol. Although this protocol is for plant genome, it can also be applied to other genome. This technique has been used for genome-wide methylation analysis and has the name of Restriction Landmark Genomic Scanning for Methylation (RLGS-M). http://www.geocities.jp/millet_kawase/RLGS/protoco... Added: Tue May 14 2002, Hits: 1508, Reviews: 0Write review