Bujalkova, Maria and Zavodna, Katarina and Krivulcik, Tomas and Ilencikova, Denisa and Wolf, Brigitte and Kovac, Michal and Karner-Hanusch, Judith and Heinimann, Karl and Marra, Giancarlo and Jiricny, Josef and Bartosova, Zdena. (2008) Multiplex SNaPshot genotyping for detecting loss of heterozygosity in the mismatch-repair genes MLH1 and MSH2 in microsatellite-unstable tumors. Clinical chemistry, 54 (11). pp. 1844-1854.
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Official URL: https://edoc.unibas.ch/63332/
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Abstract
BACKGROUND: In the workup of patients with suspected hereditary nonpolyposis colorectal cancer (HNPCC), detection of loss of heterozygosity (LOH) could help pinpoint the mismatch-repair (MMR) gene carrying the germline mutation, but analysis of microsatellite markers has proved unreliable for this purpose. We developed a simple, low-cost method based on single-nucleotide polymorphism (SNP) genotyping and capillary electrophoresis for the assessment of LOH at 2 MMR loci simultaneously. METHODS: We used the Applied Biosystems SNaPshot Multiplex Kit with meticulously selected primers to assess 14 common SNPs in MLH1 [mutL homolog 1, colon cancer, nonpolyposis type 2 (E. coli)] and MSH2 [mutS homolog 2, colon cancer, nonpolyposis type 1 (E. coli)] and optimized the protocol for DNA isolated from peripheral blood and fresh/frozen or archival microsatellite-unstable tumors from patients with confirmed (n = 42) or suspected (n = 25) HNPCC. The 42 tumors from patients with confirmed MLH1 or MSH2 germline mutations were used to validate the method's diagnostic accuracy against results obtained with DNA sequencing or multiplex ligation-dependent probe amplification. RESULTS: The SNaPshot assay provided better detection of certain SNPs than DNA sequencing. The MLH1 and MSH2 SNP marker sets were informative in 82% and 76% of the 67 cases analyzed, respectively. The new assay displayed 100% specificity for detecting LOH and predicted the location of the germline mutation in 40% of the cases (54% of those involving MLH1, 22% in MSH2). CONCLUSIONS: Our SNP-based method for detecting LOH in MLH1 and MSH2 is simple to perform with instruments available in most clinical genetics laboratories. It can be a valuable addition to protocols now used to guide mutational screening of patients with suspected HNPCC.
Faculties and Departments: | 03 Faculty of Medicine > Bereich Kinder- und Jugendheilkunde (Klinik) > Kinder- und Jugendheilkunde (UKBB) > Medizinische Genetik (Miny) 03 Faculty of Medicine > Departement Klinische Forschung > Bereich Kinder- und Jugendheilkunde (Klinik) > Kinder- und Jugendheilkunde (UKBB) > Medizinische Genetik (Miny) |
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UniBasel Contributors: | Heinimann, Karl |
Item Type: | Article, refereed |
Article Subtype: | Research Article |
Publisher: | American Association for Clinical Chemistry |
ISSN: | 0009-9147 |
e-ISSN: | 1530-8561 |
Note: | Publication type according to Uni Basel Research Database: Journal article |
Identification Number: |
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Last Modified: | 11 Aug 2020 11:55 |
Deposited On: | 11 Aug 2020 11:55 |
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