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PMID: 16670426 Published · epublish English Journal Article Research Support, Non-U.S. Gov't Validation Study

Novel method for high throughput DNA methylation marker evaluation using PNA-probe library hybridization and MALDI-TOF detection.

Nucleic acids research ·Vol. 34 ·No. 8 ·2006-05-02 ·Pages e59

Schatz P, Distler J, Berlin K, Schuster M

Abstract

The methylation of CpG dinucleotides has become a topic of great interest in cancer research, and the methylation of promoter regions of several tumor suppressor genes has been identified as a marker of tumorigenesis. Evaluation of DNA methylation markers in tumor tissue requires hundreds of samples, which must be analyzed quantitatively due to the heterogeneous composition of biological material. Therefore novel, fast and inexpensive methods for high throughput analysis are needed. Here we introduce a new assay based on peptide nucleic acid (PNA)-library hybridization and subsequent MALDI-TOF analysis. This method is multiplexable, allows the use of standard 384 well automated pipetting, and is more specific and flexible than established methods, such as microarrays and MS-SNuPE. The approach was used to evaluate three candidate colon cancer methylation markers previously identified in a microarray study. The methylation of the genes Ade-nomatous polyposis coli (APC), glycogen synthase kinase-beta-3 (GSK3beta) and eyes absent 4 (EYA4) was analyzed in 12 colon cancer and 12 normal tissues. APC and EYA4 were confirmed as being differentially methylated in colon cancer patients whereas GSK3beta did not show differential methylation.

MeSH Terms
Colorectal Neoplasms/genetics CpG Islands DNA Methylation Genes, APC Genetic Markers Genomic Library Glycogen Synthase Kinase 3/genetics Glycogen Synthase Kinase 3 beta Humans Nucleic Acid Hybridization/methods Nucleic Acid Probes Peptide Nucleic Acids/chemistry Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods Trans-Activators/genetics
Chemicals
EYA4 protein, human Genetic Markers Nucleic Acid Probes Peptide Nucleic Acids Trans-Activators GSK3B protein, human Glycogen Synthase Kinase 3 beta Glycogen Synthase Kinase 3
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Schatz Philipp
Epigenomics AG, Science Department, Kleine Präsidentenstrasse 1, D-10178 Berlin, Germany.
Distler Jürgen
Berlin Kurt
Schuster Matthias
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
1362-4962
DOI
10.1093/nar/gkl218
Published
2006-05-02
Epub
2006-00-02
Pages
e59
Language
English
Region
England
NLM ID
0411011
PMCID
PMC1456329
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/gkl218 PMC Full Text View journal details
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