# Novel Tet3 enzymes for next-generation epigenetic sequencing

## TL;DR

Researchers developed new Tet3 enzymes to map DNA methylation without harsh chemicals, improving epigenetic sequencing.

## Contribution

Novel Tet3 enzyme variants were developed for bisulfite-free epigenetic sequencing of DNA methylation.

## Key findings

- New Tet3-variants enable oxidation-based detection of 5-methylcytosine without bisulfite treatment.
- The enzymes improve accuracy and preservation of genetic material during epigenetic analysis.

## Abstract

Epigenetic regulation of gene expression is essential for cellular development and differentiation processes in higher eukaryotes. Modifications of cytosine, in particular 5-methylcytosine (5mdC), in DNA play a central role through impacting chromatin structure, repressing transposons, and regulating transcription. DNA methylation is actively installed by DNA methyltransferases and reversed through Tet-dioxygenase-mediated oxidation of 5mdC to 5-hydroxylmethylcytosine (5hmdC), 5-formylcytosine (5fdC), and 5-carboxycytosine (5cadC). It is crucial to understand the role of these epigenetic DNA modifications in cellular differentiation and developmental processes, as well as in disease state mapping and tracing of 5mdC and its oxidized forms. In bisulfite sequencing, which has been the benchmark for mapping 5mdC for the last few decades, degradation of the majority of genetic material occurs through harsh chemical treatment. Alternative sequencing methods often utilize Tet-enzyme-mediated oxidation of 5mdC to locate 5mdC and 5hmdC in genomic DNA. Herein, we report the development of novel Tet3-variants for oxidation-based bisulfite-free 5mdC- sequencing.

Epigenetic sequencing methods largely rely on oxidation of 5mdC by Tet-enzymes. We report the development and characterization of novel Tet3-variants for oxidation-based bisulfite-free 5mdC-sequencing.