DNA methylation is an heritable epigenetic mark regulated by DNA methyltransferases (DNMTs), consisting in the transfer of a methyl group onto the C5 position of a cytosine. Through this process, DNA methylation is involved in many processes like regulation of gene expression, silencing of transposable elements, genomic imprinting, and X-chromosome inactivation. It can notably explain why, even if all the cells of the organism have the same genetic information, the gene expression profile is not the same for all cell types.
In mammal somatic cells, DNA methylation is mainly located on cytosine at CpG sites and essential for many processes like normal development or cell differentiation. Global DNA hypomethylation profile with localised hypermethylation sites (like tumor suppressor gene promoters) are closely associated to cancer. Studying the DNA methylation status of key sequences can help us to understand these mechanisms.
Pyrosequencing is based on the principle of sequencing by synthesis, where nucleotides are added 1 by 1.
During the reaction, addition of dNTP is sequencially performed by cycles. The pyrosequencing instrument can monitor the real time incorporation of nucleotides through enzymatic conversion of released pyrophosphate into a proportional light signal and thus resume the exact sequence of the complementary DNA strands with pyrograms.
For the analysis of DNA methylation, required bisulfite treatment of DNA turn all non-methylated cytosines into uracils, which become thymidines after PCR amplification. So, the remaining percent of cytosine will be representative of the methylation percentage for a given position of DNA.
All our PCR primer sets are validated by PCR and qPCR, and pyrosequencing primers by pyrosequencing with strict quality control criteria to ensure you very high quality results.
Supported by bioinformatic and data mining platform, AnyGenes® provides you complete pyrosequencing services to support your scientific projects.
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