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DNA methylation is a most important epigenetic modification mode that plays crucial roles in gene expression control. DNA methylation exists in all living creatures and has several different forms. In prokaryotes, it commonly occurs on cytosine (C) in GATC and CCA/TGG. In eukaryotes, it occurs on purine, guanine, and mostly on C. Under the catalysis of MTase, the fifth carbon of the C in dinucleotide 5’-CG-3’ can be selectively modified with a methyl group to form 5-mC. The pattern of DNA methylation is inheritable and changeable. Normal expression of parental allele results in right gene expression and abnormal DNA methylation in parental allele generally causes miscellaneous cancers, genetic diseases and ageing abnormality. Therefore, research for DNA methylation has critical significance for medicine, life science and biochemistry.
Figure 1. Molecular mechanisms of DNA methylation.
Analysis of DNA methylation, DNA demethylation, and their functional effects are critical to epigenetics researchers. Methylation profiles of epigenome are used for disease identification and for research and therapeutic development. Creative Biogene can provide a comprehensive range of DNA methylation analysis to help researchers gain valuable insight into gene regulation and identify potential biomarkers. Analysis can be gene-specific or global depending on downstream applications. Depending on our client demands, we offer the following services:
Sodium bisulfite conversion of genomic DNA to differentiate and detect unmethylated versus methylated cytosines is the gold standard for DNA methylation analysis. Bisulfite conversion involves the deamination of unmodified cytosines to uracil, leaving the modified bases 5-mC and 5-hmC. This procedure can then be followed either by PCR amplification or massively parallel sequencing methods to reveal the methylation status of every cytosine in gene specific amplification or whole genome amplification.
Targeted Sanger Bisulfite Sequencing allows researchers to analyze gene regions of interest through cloning of bisulfite converted DNA fragment followed by sequencing of 10-15 clones. Amplicons are 250-450 bp in length.
Methylation-specific PCR (MSP) permits gene/sequence-specific detection of DNA methylation. Bisulfite conversion of genomic DNA followed by amplification of methylated sequences with specific primers allows the detection of methylation at specific methylation sites within genes or genomic regions.
The 5-mC DNA ELISA assay allows the quantitative analysis of global DNA methylation levels in a wide range of DNA samples. In addition, our workflow allows the analysis of a high number of samples in a short time.
Creative Biogene can provide reliable information on the methylation states of individual cytosines by effectively and efficiently preparing converted DNA for use in next generation sequencing techniques. Complete services for methylation-specific PCR and global 5-mC quantification are also available for your DNA methylation-based research. If you have any specific needs, please feel free to contact us.