Methylation Promoter PCR Kit
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Overview
· Use our Methylation Promoter PCR Kit to quickly and easily detect the methylation status of your promoter of interest, without the hassle of chemical modification with sodium bisulfate.
· Choose from our list of 40 PCR primers OR use your own PCR primers to detect methylation of promoters specific for your research
DNA methylation is a commonly occurring modification of human DNA. It involves the addition of a methyl group to cytosine residues at CpG dinucleotides, a reaction that is catalyzed by DNA methyltransferase (DNMT) enzymes. CpG dinucleotides are gathered in clusters called CpG islands, which are unequally distributed across the human genome. There are approximately 30,000 CpG islands in the genome and 50-60% of these are found within the promoter region of genes. CpG islands are primarily unmethylated in normal tissues, and the aberrant methylation of CpG islands is clearly related with disease, such as cancer.
Research shows that MeCP2 is a member of a family of proteins that selectively recognizes methylated CpGs. The binding of these proteins to DNA leads to an altered chromatin structure, which subsequently prevents the binding of transcription machinery, and thus precludes gene expression. The abnormal methylation causes transcriptional repression of numerous genes, leading to tumor growth and development. Studies of DNA methylation in cancer have uncovered new potential targets for the diagnosis, prognosis and ultimately the treatment of human cancer.
There has been a delay in the appreciation of methylation as an important epigenetic event in cancer progression. This has been due to the difficulties associated with the analysis of DNA methylation, as standard molecular biology techniques do not preserve methylation of the genomic DNA. Panomics has developed a new PCR method for analyzing methylation status of a specific promoter of interest. The method is based on MeCP2 binding, which differentiates those promoters with methylated groups from unmethylated promoters. Compared to sodium bisulphate-based conversion of methylated bases in conjunction with PCR amplification, this method is very simple and straight forward.
HOW IT WORKS
The flowchart below details how this procedure works:
Isolated genomic DNA is digested with MseI, and the resulting DNA fragments are incubated with the methylation binding protein MeCP2 (a.k.a. MBP). The methylated DNA fragments are isolated with a spin column and the amplified with promoter specific primers. Agarose gel electrophoresis is used to visualize the PCR products. The presence of a band on the gel indicates that a specific promoter is methylated in your genomic DNA sample.
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Click here for a list of all Methylation Promoter PCR Primers
DOCUMENTS
Promoter Methylation kit User Manual
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Douet, Y. et al (2008) Biochemical and Biophysical Research Communications 66-71 (PDF)
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