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Forschungsarbeit

Epi-combining - a novel method for the analysis of DNA methylation patterns at single DNA molecules

By Karina Zillner (19.09.2011)

In every single cell of our body, there is the same DNA, the same set of genes. Why does one cell develop to a neuronal cell, the other one to a liver cell? What distinguishes different type of cells?

The answer is: Cell-type specific genes must be expressed whereas other genes have to be silenced. This different, long-term interplay of active and silent genes is called epigenetic regulation. One key player is DNA methylation, which is a molecular switch that turns specific genes off when present at the start of a gene. Just like a light switch, DNA methylation is able to silence genes (figure 1). In mammalian cells, DNA methylation occurs in the context of the CpG dinucleotide at position 5 of the cytosine ring. Modifications of cytosine residues are important epigenetic regulators of genome function. Several studies showed that correct DNA methylation is crucial for development and aberrant DNA methylation is a reason for many diseases such as cancer. Dynamic DNA methylation is also tightly linked to the process of ageing. Because of this significance, considerable attempt has been made to characterize genomic DNA methylation using a wide variety of techniques.

 

Figure 1: Transcriptional silencing of gene promoters via DNA methylation

However, available approaches provide no or very limited locus-specific information about the methylation status of repetitive DNA such as ribosomal or centromeric DNA. Taking into consideration first that repetitive DNA represents more than half of the human genome, second that the majority of CpG methylation takes place in repeats, and third the importance of repeat elements in genome structure and function, there is a particularly strong need for the development of methods, which enable the genome-scale monitoring of the epigenome at the single molecule level.

To tackle this problem, the working group of Dr. Attila Németh at the University of Regensburg has combined the dynamic molecular combing technique with the simultaneous detection of DNA modifications enabling genome-scale single molecule analysis of epigenetic DNA marks. The method that we call “epi-combing” was established using in vitro methylated DNA and then applied for the characterization of higher-ordered arrangements of genomic DNA modifications on single DNA fibers (figure 2).

Figure 2: Analysis of higher-order DNA methylation patterns using "epi-combing".

We are able to show higher-ordered arrangements of the chromosomal organization of DNA methylation in single genomes at kilobase resolution. Our study initiates the locus-specific analysis of repetitive DNA modifications and opens new way for epigenomic biomarker discovery and epigenetics-based diagnostics.


Education
  • 10/2004 – 07/2009
  • Major: Biochemistry, Minor: Biophysics, degree: Diploma in Biochemistry (University of Regensburg)
  • 10/2007 – 09/2008
  • Tuition waiver for best Pre-Diploma degree of the class
  • 08/2007 – 12/2007
  • Research fellow at the Shanghai Insitute of Organic Chemistry, P.R. China
  • Since 10/2009
  • Ph.D. student in Cellular Biochemistry and Biophysics in the working group of Dr. Attila Németh (University of Regensburg)
  • Since 11/2010
  • Research fellowship of the Elite Network of Bavaria
  • Since 03/2011
  • Fellow of the Bavarian EliteAcademy (Bayerische EliteAkademie)

Publications
  • Gaussia-luciferase as a sensitive reporter gene for monitoring promoter activity in the nucleus of the green alga Chlamydomonas reinhardtii, Mol Genet Genomics, 280, 2, Seiten 153-162 (May 2008)
  • Microscale Thermophoresis as a sensitive method to quantify protein –nucleic acid interactions in solution, Methods in Molecular Biology – Functional Genomics, Springer Verlag (in press, December 2011)

Research interests
  • Nuclear Architecture
  • Epigenetics