Chemical assisted enrichment of 5-carboxycytosine that also allows for DNA sequencing at single base resolution.

ended

5-carboxycytosine (5caC) has been robustly identified in mammalian DNA. It is known that this DNA modification plays a role in epigenetic demethylation processes. If 5caC has other epigenetically relevant functions is so far unknown. Due to its low abundance it is technically challenging to study this modification in biological samples. Identification of specific readers, recognition by the RNA polymerase II elongation complex, the presence in specific genomic loci as well as changing levels during differentiation of mouse embryonic stem cells (mESCs) point towards an important biological function. A chemical tool that would clarify the role of 5caC in mammalian biology is so far missing. Since DNA modifications play a significant role in human diseases, understanding their functions offer new opportunities for novel treatment strategies. Chemical tools have advanced the field enormously in detecting and analysing newly discovered modified bases by rationally designed chemistry for specific labelling of a given modification. Chemical enrichment of DNA fragments enables mapping of the bases at 200-400 base pair resolution and has been used for a genome-wide mapping of several DNA modifications. To obtain single base resolution a separate modification specific sequencing method needs to be applied after enrichment. To avoid a multistep procedure I will develop a chemical enrichment method for 5caC that is directly coupled to single base resolution sequencing. In combination this provides a powerful chemical tool that has far reaching impact for other researchers in the field and finally enables scientific progress on 5caC.