Analysis of DNA Methylation in FFPE Tissues Using the MethyLight Technology
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Novel biomarkers are sought after by mining DNA extracted from formalin-fixed, paraffin-embedded (FFPE) tissues. Such tissues offer the great advantage of often having complete clinical data (including survival), as well as the tissues are amenable for laser microdissection targeting specific tissue areas. Downstream analysis of such DNA includes mutational screens and methylation profiling. Screening for mutations by sequencing requires a significant amount of DNA for PCR and cycle sequencing. This is self-inhibitory if the gene screened has a large number of exons. Profiling DNA methylation using the MethyLight technology circumvents this problem and allows for the mining of several biomarkers from DNA extracted from a single microscope slide of the tissue of interest. We describe in this chapter a detailed protocol for MethyLight and its use in the determination of CpG Island Methylator Phenotype status in FFPE colorectal cancer samples.
Key wordsPCR MethyLight FFPE DNA Methylation CIMP PMR
This work is supported by Kuwait Foundation for the Advancement of Sciences grant number 2006-1302-07, Kuwait University Grant number MG02/08, and Research Core Facility (RCF) grant number GM 01/01and GM 01/05.
- 2.Gagnon, J. F., Sanschagrin, F., Jacob, S., Tremblay, A. A., Provencher, L., Robert, J., et al. (2009) Quantitative DNA methylation analysis of laser capture microdissected formalin-fixed and paraffin-embedded tissues. Exp Mol Pathol. 8, 8.Google Scholar