Multiplex SNaPshot Genotyping for Detecting Loss of Heterozygosity in the Mismatch-Repair Genes MLH1 and MSH2 in Microsatellite-Unstable Tumors
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 d...
Gespeichert in:
| Veröffentlicht in: | Clinical chemistry (Baltimore, Md.) Md.), 2008-11, Vol.54 (11), p.1844-1854 |
|---|---|
| Hauptverfasser: | , , , , , , , , , , |
| Format: | Artikel |
| Sprache: | eng |
| Schlagworte: | |
| Online-Zugang: | Volltext |
| Tags: |
Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
|
| container_end_page | 1854 |
|---|---|
| container_issue | 11 |
| container_start_page | 1844 |
| container_title | Clinical chemistry (Baltimore, Md.) |
| container_volume | 54 |
| creator | Bujalkova, Maria Zavodna, Katarina Krivulcik, Tomas Ilencikova, Denisa Wolf, Brigitte Kovac, Michal Karner-Hanusch, Judith Heinimann, Karl Marra, Giancarlo Jiricny, Josef Bartosova, Zdena |
| description | 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.
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.
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).
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. |
| doi_str_mv | 10.1373/clinchem.2008.108902 |
| format | Article |
| fullrecord | <record><control><sourceid>proquest_cross</sourceid><recordid>TN_cdi_proquest_miscellaneous_69717634</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>1592064371</sourcerecordid><originalsourceid>FETCH-LOGICAL-c440t-c4dc04004ddd7febc476fa820ef9727495526f96fb0ca9bbbb717a741ceac6e83</originalsourceid><addsrcrecordid>eNpdkdGK1DAUhoMo7rj6BiJBUK86JmnapJey6o4wVXF3r0MmPZ1maZuapIzjS_jKpsyoYC4STvjOzzl8CD2nZE1zkb81vR1NB8OaESLXlMiKsAdoRYucZLIo6UO0IoRUWUW5uEBPQrhPJReyfIwuqBSC5ZSs0K967qOdeviBbz7rr6FzEV_D6OJxsuMet87j9xDBxKXauhCwa_Em_Xj387h3wcYjtiOOHeDahkFH02XfYNLWLzEQcL3dUKzHBtc3G7agtTXeBR2h722E7G4MUe96wLfz4Hx4ih61ug_w7PxeoruPH26vNtn2y_Wnq3fbzHBOYrobQ3jap2ka0cLOcFG2WjICbSWY4FVRsLKtynZHjK526QgqtODUgDYlyPwSvT7lTt59nyFENdhg0kx6BDcHVVapocx5Al_-B9672Y9pNsVoXsmCVSxB_AQtqwUPrZq8HbQ_KkrUYkv9saUWW-pkK7W9OGfPuwGaf01nPQl4dQZ0MLpvvR6NDX85RiSVkpSJe3PiOrvvDtaDSir6PsVSdTgcCq4oTamc578BkOiupA</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>213985292</pqid></control><display><type>article</type><title>Multiplex SNaPshot Genotyping for Detecting Loss of Heterozygosity in the Mismatch-Repair Genes MLH1 and MSH2 in Microsatellite-Unstable Tumors</title><source>Oxford University Press Journals All Titles (1996-Current)</source><source>MEDLINE</source><creator>Bujalkova, Maria ; Zavodna, Katarina ; Krivulcik, Tomas ; Ilencikova, Denisa ; Wolf, Brigitte ; Kovac, Michal ; Karner-Hanusch, Judith ; Heinimann, Karl ; Marra, Giancarlo ; Jiricny, Josef ; Bartosova, Zdena</creator><creatorcontrib>Bujalkova, Maria ; Zavodna, Katarina ; Krivulcik, Tomas ; Ilencikova, Denisa ; Wolf, Brigitte ; Kovac, Michal ; Karner-Hanusch, Judith ; Heinimann, Karl ; Marra, Giancarlo ; Jiricny, Josef ; Bartosova, Zdena</creatorcontrib><description>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.
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.
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).
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.</description><identifier>ISSN: 0009-9147</identifier><identifier>EISSN: 1530-8561</identifier><identifier>DOI: 10.1373/clinchem.2008.108902</identifier><identifier>PMID: 18772310</identifier><identifier>CODEN: CLCHAU</identifier><language>eng</language><publisher>Washington, DC: Am Assoc Clin Chem</publisher><subject>Adaptor Proteins, Signal Transducing - genetics ; Analytical, structural and metabolic biochemistry ; Base Pair Mismatch - genetics ; Base Sequence ; Biological and medical sciences ; Cancer ; Colorectal carcinoma ; Colorectal Neoplasms, Hereditary Nonpolyposis - genetics ; Deoxyribonucleic acid ; DNA ; DNA Primers ; DNA Repair - genetics ; E coli ; Fundamental and applied biological sciences. Psychology ; Genes ; Genetic testing ; Genetics ; Genotype ; Humans ; Investigative techniques, diagnostic techniques (general aspects) ; Loss of Heterozygosity ; Mass spectrometry ; Medical sciences ; Methods ; Mutation ; MutL Protein Homolog 1 ; MutS Homolog 2 Protein - genetics ; Nuclear Proteins - genetics ; Polymerase Chain Reaction ; Polymorphism, Single Nucleotide ; Roles ; Sensitivity and Specificity ; Studies</subject><ispartof>Clinical chemistry (Baltimore, Md.), 2008-11, Vol.54 (11), p.1844-1854</ispartof><rights>2008 INIST-CNRS</rights><rights>Copyright American Association for Clinical Chemistry Nov 2008</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c440t-c4dc04004ddd7febc476fa820ef9727495526f96fb0ca9bbbb717a741ceac6e83</citedby><cites>FETCH-LOGICAL-c440t-c4dc04004ddd7febc476fa820ef9727495526f96fb0ca9bbbb717a741ceac6e83</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=20818806$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/18772310$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Bujalkova, Maria</creatorcontrib><creatorcontrib>Zavodna, Katarina</creatorcontrib><creatorcontrib>Krivulcik, Tomas</creatorcontrib><creatorcontrib>Ilencikova, Denisa</creatorcontrib><creatorcontrib>Wolf, Brigitte</creatorcontrib><creatorcontrib>Kovac, Michal</creatorcontrib><creatorcontrib>Karner-Hanusch, Judith</creatorcontrib><creatorcontrib>Heinimann, Karl</creatorcontrib><creatorcontrib>Marra, Giancarlo</creatorcontrib><creatorcontrib>Jiricny, Josef</creatorcontrib><creatorcontrib>Bartosova, Zdena</creatorcontrib><title>Multiplex SNaPshot Genotyping for Detecting Loss of Heterozygosity in the Mismatch-Repair Genes MLH1 and MSH2 in Microsatellite-Unstable Tumors</title><title>Clinical chemistry (Baltimore, Md.)</title><addtitle>Clin Chem</addtitle><description>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.
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.
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).
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.</description><subject>Adaptor Proteins, Signal Transducing - genetics</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>Base Pair Mismatch - genetics</subject><subject>Base Sequence</subject><subject>Biological and medical sciences</subject><subject>Cancer</subject><subject>Colorectal carcinoma</subject><subject>Colorectal Neoplasms, Hereditary Nonpolyposis - genetics</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>DNA Primers</subject><subject>DNA Repair - genetics</subject><subject>E coli</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Genes</subject><subject>Genetic testing</subject><subject>Genetics</subject><subject>Genotype</subject><subject>Humans</subject><subject>Investigative techniques, diagnostic techniques (general aspects)</subject><subject>Loss of Heterozygosity</subject><subject>Mass spectrometry</subject><subject>Medical sciences</subject><subject>Methods</subject><subject>Mutation</subject><subject>MutL Protein Homolog 1</subject><subject>MutS Homolog 2 Protein - genetics</subject><subject>Nuclear Proteins - genetics</subject><subject>Polymerase Chain Reaction</subject><subject>Polymorphism, Single Nucleotide</subject><subject>Roles</subject><subject>Sensitivity and Specificity</subject><subject>Studies</subject><issn>0009-9147</issn><issn>1530-8561</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2008</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>BENPR</sourceid><recordid>eNpdkdGK1DAUhoMo7rj6BiJBUK86JmnapJey6o4wVXF3r0MmPZ1maZuapIzjS_jKpsyoYC4STvjOzzl8CD2nZE1zkb81vR1NB8OaESLXlMiKsAdoRYucZLIo6UO0IoRUWUW5uEBPQrhPJReyfIwuqBSC5ZSs0K967qOdeviBbz7rr6FzEV_D6OJxsuMet87j9xDBxKXauhCwa_Em_Xj387h3wcYjtiOOHeDahkFH02XfYNLWLzEQcL3dUKzHBtc3G7agtTXeBR2h722E7G4MUe96wLfz4Hx4ih61ug_w7PxeoruPH26vNtn2y_Wnq3fbzHBOYrobQ3jap2ka0cLOcFG2WjICbSWY4FVRsLKtynZHjK526QgqtODUgDYlyPwSvT7lTt59nyFENdhg0kx6BDcHVVapocx5Al_-B9672Y9pNsVoXsmCVSxB_AQtqwUPrZq8HbQ_KkrUYkv9saUWW-pkK7W9OGfPuwGaf01nPQl4dQZ0MLpvvR6NDX85RiSVkpSJe3PiOrvvDtaDSir6PsVSdTgcCq4oTamc578BkOiupA</recordid><startdate>20081101</startdate><enddate>20081101</enddate><creator>Bujalkova, Maria</creator><creator>Zavodna, Katarina</creator><creator>Krivulcik, Tomas</creator><creator>Ilencikova, Denisa</creator><creator>Wolf, Brigitte</creator><creator>Kovac, Michal</creator><creator>Karner-Hanusch, Judith</creator><creator>Heinimann, Karl</creator><creator>Marra, Giancarlo</creator><creator>Jiricny, Josef</creator><creator>Bartosova, Zdena</creator><general>Am Assoc Clin Chem</general><general>American Association for Clinical Chemistry</general><general>Oxford University Press</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>3V.</scope><scope>4U-</scope><scope>7QO</scope><scope>7RV</scope><scope>7TM</scope><scope>7U7</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>88I</scope><scope>8AO</scope><scope>8C1</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABJCF</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>BKSAR</scope><scope>C1K</scope><scope>CCPQU</scope><scope>D1I</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>KB.</scope><scope>KB0</scope><scope>M0S</scope><scope>M1P</scope><scope>M2P</scope><scope>NAPCQ</scope><scope>P64</scope><scope>PCBAR</scope><scope>PDBOC</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>Q9U</scope><scope>RC3</scope><scope>S0X</scope><scope>7X8</scope></search><sort><creationdate>20081101</creationdate><title>Multiplex SNaPshot Genotyping for Detecting Loss of Heterozygosity in the Mismatch-Repair Genes MLH1 and MSH2 in Microsatellite-Unstable Tumors</title><author>Bujalkova, Maria ; Zavodna, Katarina ; Krivulcik, Tomas ; Ilencikova, Denisa ; Wolf, Brigitte ; Kovac, Michal ; Karner-Hanusch, Judith ; Heinimann, Karl ; Marra, Giancarlo ; Jiricny, Josef ; Bartosova, Zdena</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c440t-c4dc04004ddd7febc476fa820ef9727495526f96fb0ca9bbbb717a741ceac6e83</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2008</creationdate><topic>Adaptor Proteins, Signal Transducing - genetics</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>Base Pair Mismatch - genetics</topic><topic>Base Sequence</topic><topic>Biological and medical sciences</topic><topic>Cancer</topic><topic>Colorectal carcinoma</topic><topic>Colorectal Neoplasms, Hereditary Nonpolyposis - genetics</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>DNA Primers</topic><topic>DNA Repair - genetics</topic><topic>E coli</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Genes</topic><topic>Genetic testing</topic><topic>Genetics</topic><topic>Genotype</topic><topic>Humans</topic><topic>Investigative techniques, diagnostic techniques (general aspects)</topic><topic>Loss of Heterozygosity</topic><topic>Mass spectrometry</topic><topic>Medical sciences</topic><topic>Methods</topic><topic>Mutation</topic><topic>MutL Protein Homolog 1</topic><topic>MutS Homolog 2 Protein - genetics</topic><topic>Nuclear Proteins - genetics</topic><topic>Polymerase Chain Reaction</topic><topic>Polymorphism, Single Nucleotide</topic><topic>Roles</topic><topic>Sensitivity and Specificity</topic><topic>Studies</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Bujalkova, Maria</creatorcontrib><creatorcontrib>Zavodna, Katarina</creatorcontrib><creatorcontrib>Krivulcik, Tomas</creatorcontrib><creatorcontrib>Ilencikova, Denisa</creatorcontrib><creatorcontrib>Wolf, Brigitte</creatorcontrib><creatorcontrib>Kovac, Michal</creatorcontrib><creatorcontrib>Karner-Hanusch, Judith</creatorcontrib><creatorcontrib>Heinimann, Karl</creatorcontrib><creatorcontrib>Marra, Giancarlo</creatorcontrib><creatorcontrib>Jiricny, Josef</creatorcontrib><creatorcontrib>Bartosova, Zdena</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>University Readers</collection><collection>Biotechnology Research Abstracts</collection><collection>Nursing & Allied Health Database</collection><collection>Nucleic Acids Abstracts</collection><collection>Toxicology Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>Science Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>Public Health Database</collection><collection>Technology Research Database</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Technology Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>Materials Science & Engineering Collection</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest One Sustainability</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>ProQuest Central</collection><collection>Technology Collection</collection><collection>Natural Science Collection</collection><collection>Earth, Atmospheric & Aquatic Science Collection</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ProQuest One Community College</collection><collection>ProQuest Materials Science Collection</collection><collection>ProQuest Central Korea</collection><collection>Engineering Research Database</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Materials Science Database</collection><collection>Nursing & Allied Health Database (Alumni Edition)</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Science Database</collection><collection>Nursing & Allied Health Premium</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Earth, Atmospheric & Aquatic Science Database</collection><collection>Materials Science Collection</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central Basic</collection><collection>Genetics Abstracts</collection><collection>SIRS Editorial</collection><collection>MEDLINE - Academic</collection><jtitle>Clinical chemistry (Baltimore, Md.)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Bujalkova, Maria</au><au>Zavodna, Katarina</au><au>Krivulcik, Tomas</au><au>Ilencikova, Denisa</au><au>Wolf, Brigitte</au><au>Kovac, Michal</au><au>Karner-Hanusch, Judith</au><au>Heinimann, Karl</au><au>Marra, Giancarlo</au><au>Jiricny, Josef</au><au>Bartosova, Zdena</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Multiplex SNaPshot Genotyping for Detecting Loss of Heterozygosity in the Mismatch-Repair Genes MLH1 and MSH2 in Microsatellite-Unstable Tumors</atitle><jtitle>Clinical chemistry (Baltimore, Md.)</jtitle><addtitle>Clin Chem</addtitle><date>2008-11-01</date><risdate>2008</risdate><volume>54</volume><issue>11</issue><spage>1844</spage><epage>1854</epage><pages>1844-1854</pages><issn>0009-9147</issn><eissn>1530-8561</eissn><coden>CLCHAU</coden><abstract>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.
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.
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).
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.</abstract><cop>Washington, DC</cop><pub>Am Assoc Clin Chem</pub><pmid>18772310</pmid><doi>10.1373/clinchem.2008.108902</doi><tpages>11</tpages><oa>free_for_read</oa></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0009-9147 |
| ispartof | Clinical chemistry (Baltimore, Md.), 2008-11, Vol.54 (11), p.1844-1854 |
| issn | 0009-9147 1530-8561 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_69717634 |
| source | Oxford University Press Journals All Titles (1996-Current); MEDLINE |
| subjects | Adaptor Proteins, Signal Transducing - genetics Analytical, structural and metabolic biochemistry Base Pair Mismatch - genetics Base Sequence Biological and medical sciences Cancer Colorectal carcinoma Colorectal Neoplasms, Hereditary Nonpolyposis - genetics Deoxyribonucleic acid DNA DNA Primers DNA Repair - genetics E coli Fundamental and applied biological sciences. Psychology Genes Genetic testing Genetics Genotype Humans Investigative techniques, diagnostic techniques (general aspects) Loss of Heterozygosity Mass spectrometry Medical sciences Methods Mutation MutL Protein Homolog 1 MutS Homolog 2 Protein - genetics Nuclear Proteins - genetics Polymerase Chain Reaction Polymorphism, Single Nucleotide Roles Sensitivity and Specificity Studies |
| title | Multiplex SNaPshot Genotyping for Detecting Loss of Heterozygosity in the Mismatch-Repair Genes MLH1 and MSH2 in Microsatellite-Unstable Tumors |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-02-10T11%3A57%3A10IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-proquest_cross&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Multiplex%20SNaPshot%20Genotyping%20for%20Detecting%20Loss%20of%20Heterozygosity%20in%20the%20Mismatch-Repair%20Genes%20MLH1%20and%20MSH2%20in%20Microsatellite-Unstable%20Tumors&rft.jtitle=Clinical%20chemistry%20(Baltimore,%20Md.)&rft.au=Bujalkova,%20Maria&rft.date=2008-11-01&rft.volume=54&rft.issue=11&rft.spage=1844&rft.epage=1854&rft.pages=1844-1854&rft.issn=0009-9147&rft.eissn=1530-8561&rft.coden=CLCHAU&rft_id=info:doi/10.1373/clinchem.2008.108902&rft_dat=%3Cproquest_cross%3E1592064371%3C/proquest_cross%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=213985292&rft_id=info:pmid/18772310&rfr_iscdi=true |