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Advances in DNA methylation research have led to the discovery of multiple cytosine methylation variants that are derived from the oxidation of 5-methylcytosine through the activity of the TET family of enzymes. The modifications include 5-hydroxymethylcytosine (5-hmC) and, more recently, 5-formylcytosine (5-fC) & 5-carboxylcytosine (5-caC). Given the newness of this field the biological relevance of these modifications is not fully understood, however, new tools and emerging technologies will enable researchers to discover the importance of these modifications in cellular biology.
Active Motif’s 5-fC and 5-caC Profiling Services are based on DNA immunoprecipitation using antibodies that specifically recognize each modification. Our immunoprecipitation protocols have been optimized to overcome the challenges presented by the low levels of 5-fC and 5-caC that are present in the genome compared to 5-methylcytosine levels. Following enrichment, DNA is sequenced using Illumina Next-Gen sequencing technology, resulting in genome-wide enrichment profiles of 5-formyl and 5-carboxylcytosines.
The 5-fC and 5-caC Profiling Service includes:
- Preparation of the DNA sample*
- Immunoprecipitation using 5-fC or 5-caC antibodies
- Amplification of the enriched DNA sample
- Next-Gen sequencing
- Analysis of the data and delivery to the customer
Figure 1: Genome-wide profiling of the DNA variants 5-hmC, 5-fC and 5-caC.