Cells have a broad functional and morphological diversity due to differential gene expression. Research in epigenetics combines the study of inheritable, phenotypical changes in the gene expression pattern of a specific cell type that are not caused by a transformed nucleotide sequence of the genetic code itself. Epigenetic marks are represented by a variety of molecular mechanisms including DNA methylation. Alterations of DNA methylation play a crucial role in the onset of diseases like cancer. Many DNA methylation-based biomarkers have been evaluated and the analysis of epigenetic alterations is a promising tool for disease diagnostics, prognostics, and prediction of drug response. In future, this will allow to adapt therapies to a person, which will increase the chance for successful treatments, minimizing side-effects of chemotherapy and administration of ineffective drugs and thus, prevent the onset of follow-up problems that are associated with these events. Thus, cost-effective but robust means that allow the analysis of DNA methylation-based biomarkers are of urgent need. Several methods for analysis of these biomarkers are employed. However, those that have the required resolution are laborious, time-consuming, and error-prone and thus, prevent broad applications of DNA methylation profiling in clinical diagnostics. The aim of this project is to overcome the barriers that prohibit using DNA methylation profiling in broad clinical applications for diagnostics, prognostics, and prediction of drug response. The objectives will be reached by a multidisciplinary systemic approach harnessing the power of organic synthesis (i.e. new synthetic modified nucleotides), biochemical and structural enzyme studies, and directed evolution of DNA polymerases tailored for new replication systems for epigenetics. The evolved replication systems will be superior to known techniques by superseding the bottle necks of current approaches paving the way for broad applications.