Prevention of DNA Fragmentation Improves DNA Methylation Analysis

DNA methylation is the most intensively studied epigenetic modification and describes the conversion of cytosine to 5-methylcytosine. The importance of this phenomenon is that aberrant promoter hypermethylation is frequently associated with gene silencing and is a common occurrence in cancer. The most critical step for correct determination of DNA methylation status is the complete conversion of unmethylated cytosines to uracil. Conversion of cytosines is achieved by bisulfite treatment of DNA at high temperature and low pH values. These harsh conditions lead to a high degree of DNA fragmentation and subsequent loss of DNA. Due to this loss of DNA, common bisulfite procedures usually require high amounts of input DNA — often leading to low DNA yield, highly fragmented DNA, and irreproducible conversion rates. This clearly hampers methylation analysis when using small amounts of input DNA (e.g., DNA isolated from microdissected biopsies, formalin-fixed paraffin-embedded tissues, or other small and precious sample materials).

Here, we present data on the following topics:

  • Reliable methylation analysis through high bisulfite conversion efficiency
  • Innovative DNA protection mechanism resulting in increased assay sensitivity
  • Long-term study on DNA integrity and conversion stability
  • Cancer research methylation analysis using Pyrosequencing
  • Methylation profiling of FFPE samples using a specialized protocol
  • EpiTect technology for reliable and highly sensitive epigenetics research

Dr. Thorsten Traeger

Thorsten Traeger is a member of the Epigenetics application group where he is involved in the development of the EpiTect product line. Dr. Traeger received his PhD from the Ruhr-University Bochum, Germany and joined QIAGEN 9 years ago, starting in the sequencing department before switching to the Modification and Amplification group, where he is responsible for the development of products and assays for real-time PCR and also for epigenetic applications. Currently, Dr. Traeger is an Associate Director, Project Management R&D in the Modification and Amplification Group at QIAGEN.