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PubMed · 16101135

A new method for optimizing multiplex DNA microsatellite analysis in low quality archival specimens.

Irina Drobinskaya·Helmut Erich Gabbert·Gabriela Moeslein·Wolfram Mueller
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Abstract

BACKGROUND: Testing microsatellite instability seems to be a useful tool for the initial screening of putative non-polyposis colorectal cancer (HNPCC), preceding analysis of germ-line mutations of DNA mismatch repair genes. However, diagnosis of microsatellite instability becomes complicated when highly-damaged DNA from formalin-fixed paraffin-embedded tissue specimens has to be investigated. MATERIALS AND METHODS: A new methodical approach was established based on special multiplex PCR regimes (e.g., on touch-up cycling conditions), allowing both sufficient, as well as specific, amplification of the Bethesda reference panel loci with low quality DNA as template. RESULTS: By applying our new method, microsatellite instability could be analyzed successfully in 75 out of 84 investigated tumors (89%), whereas, by using standard PCR protocols, microsatellite analysis failed in 36% of the investigated cases. CONCLUSION: Our new methodical approach should be recommended for the use of archival material since it allows an efficient and accurate amplification of the Bethesda marker fragments and is less dependent on the DNA quality.

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Adaptor Proteins, Signal TransducingAllelic ImbalanceCarrier ProteinsColorectal Neoplasms, Hereditary NonpolyposisDNA DamageDNA, NeoplasmDNA-Binding ProteinsFormaldehydeHumansImmunohistochemistryMicrosatellite RepeatsMutL Protein Homolog 1MutS Homolog 2 ProteinNeoplasm ProteinsNuclear ProteinsParaffin EmbeddingPolymerase Chain ReactionProto-Oncogene ProteinsTissue FixationJournal ArticleResearch Support, Non-U.S. Gov't

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Irina Drobinskaya, Helmut Erich Gabbert, Gabriela Moeslein, Wolfram Mueller. A new method for optimizing multiplex DNA microsatellite analysis in low quality archival specimens.. https://pubmed.ncbi.nlm.nih.gov/16101135/

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