Digital PCR for discriminating mosaic deletions and for determining proportion of tumor cells in specimen
Mosaicism, presence of a genetic feature in only a subpopulation of cells, is frequent in de novo genetic diseases. Among large deletions covering the NF1 tumor suppressor gene, the frequency of mosaicism can be as high as 40% in de novo patients. In this study, we demonstrate the high potential of...
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| Veröffentlicht in: | European journal of human genetics : EJHG 2016-11, Vol.24 (11), p.1644-1648 |
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| description | Mosaicism, presence of a genetic feature in only a subpopulation of cells, is frequent in de novo genetic diseases. Among large deletions covering the NF1 tumor suppressor gene, the frequency of mosaicism can be as high as 40% in de novo patients. In this study, we demonstrate the high potential of digital PCR in detecting large NF1 deletions and in discriminating mosaic cases. By simultaneously assessing the NF1 gene and a reference gene RPP30, deletions could be unambiguously distinguished from non-deletion samples. Performing the same assay for mixed samples from a DNA with a deletion and a non-deletion DNA, a highly significant linear relation was obtained between the set-up ratio of the two samples and the measured ratio of NF1/RPP30 (P |
| doi_str_mv | 10.1038/ejhg.2016.56 |
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Among large deletions covering the NF1 tumor suppressor gene, the frequency of mosaicism can be as high as 40% in de novo patients. In this study, we demonstrate the high potential of digital PCR in detecting large NF1 deletions and in discriminating mosaic cases. By simultaneously assessing the NF1 gene and a reference gene RPP30, deletions could be unambiguously distinguished from non-deletion samples. Performing the same assay for mixed samples from a DNA with a deletion and a non-deletion DNA, a highly significant linear relation was obtained between the set-up ratio of the two samples and the measured ratio of NF1/RPP30 (P<0.0001), suggesting the high potential of digital PCR in discriminating mosaic deletions. Furthermore, digital PCR detects NF1 allele loss in a tumor specimen that was not detected by loss of heterozygosity analysis using polymorphic markers due to high content of non-tumor cells. Based on the measured ratio of NF1/RPP30, the proportion of the tumor cells in this specimen could be calculated as 25%. Our results demonstrate that dual-probe digital PCR is a simple and effective method for detecting deletions and for discriminating mosaic deletions. Furthermore, this method is sensitive for assigning somatic allele loss in tumor specimen and enables determining proportion of tumor cells.</description><identifier>ISSN: 1018-4813</identifier><identifier>EISSN: 1476-5438</identifier><identifier>DOI: 10.1038/ejhg.2016.56</identifier><identifier>PMID: 27273132</identifier><language>eng</language><publisher>England: Nature Publishing Group</publisher><subject>Alleles ; Cell Separation - methods ; Clonal deletion ; Deoxyribonucleic acid ; DNA ; DNA, Neoplasm - chemistry ; Gene Deletion ; Genetic testing ; Genetics ; Heterozygosity ; Humans ; Laboratories ; Loss of heterozygosity ; Maxillofacial surgery ; Mosaicism ; Neurofibromatosis 1 - genetics ; Neurofibromatosis 1 - pathology ; Neurofibromin 1 ; Neurofibromin 1 - genetics ; Patients ; Polymerase chain reaction ; Ratios ; Real-Time Polymerase Chain Reaction - methods ; Sensitivity and Specificity ; Short Report ; Tumor cells ; Tumor suppressor genes ; Tumors</subject><ispartof>European journal of human genetics : EJHG, 2016-11, Vol.24 (11), p.1644-1648</ispartof><rights>Copyright Nature Publishing Group Nov 2016</rights><rights>Copyright © 2016 Macmillan Publishers Limited 2016 Macmillan Publishers Limited</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c511t-e78bc03b534dadc2b243dc68156441763edbb460d9c543168caecf76d54cd2883</citedby><cites>FETCH-LOGICAL-c511t-e78bc03b534dadc2b243dc68156441763edbb460d9c543168caecf76d54cd2883</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5110055/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5110055/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,314,727,780,784,885,27924,27925,53791,53793</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/27273132$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Kluwe, Lan</creatorcontrib><title>Digital PCR for discriminating mosaic deletions and for determining proportion of tumor cells in specimen</title><title>European journal of human genetics : EJHG</title><addtitle>Eur J Hum Genet</addtitle><description>Mosaicism, presence of a genetic feature in only a subpopulation of cells, is frequent in de novo genetic diseases. Among large deletions covering the NF1 tumor suppressor gene, the frequency of mosaicism can be as high as 40% in de novo patients. In this study, we demonstrate the high potential of digital PCR in detecting large NF1 deletions and in discriminating mosaic cases. By simultaneously assessing the NF1 gene and a reference gene RPP30, deletions could be unambiguously distinguished from non-deletion samples. Performing the same assay for mixed samples from a DNA with a deletion and a non-deletion DNA, a highly significant linear relation was obtained between the set-up ratio of the two samples and the measured ratio of NF1/RPP30 (P<0.0001), suggesting the high potential of digital PCR in discriminating mosaic deletions. Furthermore, digital PCR detects NF1 allele loss in a tumor specimen that was not detected by loss of heterozygosity analysis using polymorphic markers due to high content of non-tumor cells. Based on the measured ratio of NF1/RPP30, the proportion of the tumor cells in this specimen could be calculated as 25%. Our results demonstrate that dual-probe digital PCR is a simple and effective method for detecting deletions and for discriminating mosaic deletions. Furthermore, this method is sensitive for assigning somatic allele loss in tumor specimen and enables determining proportion of tumor cells.</description><subject>Alleles</subject><subject>Cell Separation - methods</subject><subject>Clonal deletion</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>DNA, Neoplasm - chemistry</subject><subject>Gene Deletion</subject><subject>Genetic testing</subject><subject>Genetics</subject><subject>Heterozygosity</subject><subject>Humans</subject><subject>Laboratories</subject><subject>Loss of heterozygosity</subject><subject>Maxillofacial surgery</subject><subject>Mosaicism</subject><subject>Neurofibromatosis 1 - genetics</subject><subject>Neurofibromatosis 1 - pathology</subject><subject>Neurofibromin 1</subject><subject>Neurofibromin 1 - genetics</subject><subject>Patients</subject><subject>Polymerase chain reaction</subject><subject>Ratios</subject><subject>Real-Time Polymerase Chain Reaction - methods</subject><subject>Sensitivity and Specificity</subject><subject>Short Report</subject><subject>Tumor cells</subject><subject>Tumor suppressor genes</subject><subject>Tumors</subject><issn>1018-4813</issn><issn>1476-5438</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2016</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNqNkc1rFDEAxUOxtHX15lkCXjw4a74nexHKttVCwVL0HDJJZptlJhmTGcH_vgnbFvXUUwLvx0veewC8w2iNEZWf3f5-tyYIizUXR-AMs1Y0nFH5qtwRlg2TmJ6C1znvESpii0_AKWlJSzElZ8Bf-J2f9QBvt3ewjwlan03yow969mEHx5i1N9C6wc0-hgx1sAfOzS4VrEJTilNMVYexh_MyFt24YcjQB5gnZ_zowhtw3Oshu7eP5wr8vLr8sf3W3Hz_er09v2kMx3huXCs7g2jHKbPaGtIRRq0REnPBGG4FdbbrmEB2Y0pKLKTRzvStsJwZS6SkK_Dl4Dst3eiscWFOelBTCaXTHxW1V_8qwd-rXfytyvMIcV4MPj4apPhrcXlWY-mkxNHBxSUrLGkpD2_oS1AixIaRtqIf_kP3cUmhNFEpKRmu4Vbg04EyKeacXP_8b4xUnVvVuVWdW_GKv_876zP8tC99ANEtp5s</recordid><startdate>20161101</startdate><enddate>20161101</enddate><creator>Kluwe, Lan</creator><general>Nature Publishing Group</general><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>7X7</scope><scope>7XB</scope><scope>88A</scope><scope>88E</scope><scope>8AO</scope><scope>8FD</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M7P</scope><scope>P64</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20161101</creationdate><title>Digital PCR for discriminating mosaic deletions and for determining proportion of tumor cells in specimen</title><author>Kluwe, Lan</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c511t-e78bc03b534dadc2b243dc68156441763edbb460d9c543168caecf76d54cd2883</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2016</creationdate><topic>Alleles</topic><topic>Cell Separation - methods</topic><topic>Clonal deletion</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>DNA, Neoplasm - chemistry</topic><topic>Gene Deletion</topic><topic>Genetic testing</topic><topic>Genetics</topic><topic>Heterozygosity</topic><topic>Humans</topic><topic>Laboratories</topic><topic>Loss of heterozygosity</topic><topic>Maxillofacial surgery</topic><topic>Mosaicism</topic><topic>Neurofibromatosis 1 - genetics</topic><topic>Neurofibromatosis 1 - pathology</topic><topic>Neurofibromin 1</topic><topic>Neurofibromin 1 - genetics</topic><topic>Patients</topic><topic>Polymerase chain reaction</topic><topic>Ratios</topic><topic>Real-Time Polymerase Chain Reaction - methods</topic><topic>Sensitivity and Specificity</topic><topic>Short Report</topic><topic>Tumor cells</topic><topic>Tumor suppressor genes</topic><topic>Tumors</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kluwe, Lan</creatorcontrib><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>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Biology Database (Alumni Edition)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>Technology Research Database</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</collection><collection>ProQuest One Community College</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>ProQuest Biological Science Collection</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Biological Science Database</collection><collection>Biotechnology and BioEngineering Abstracts</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 China</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>European journal of human genetics : EJHG</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Kluwe, Lan</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Digital PCR for discriminating mosaic deletions and for determining proportion of tumor cells in specimen</atitle><jtitle>European journal of human genetics : EJHG</jtitle><addtitle>Eur J Hum Genet</addtitle><date>2016-11-01</date><risdate>2016</risdate><volume>24</volume><issue>11</issue><spage>1644</spage><epage>1648</epage><pages>1644-1648</pages><issn>1018-4813</issn><eissn>1476-5438</eissn><abstract>Mosaicism, presence of a genetic feature in only a subpopulation of cells, is frequent in de novo genetic diseases. Among large deletions covering the NF1 tumor suppressor gene, the frequency of mosaicism can be as high as 40% in de novo patients. In this study, we demonstrate the high potential of digital PCR in detecting large NF1 deletions and in discriminating mosaic cases. By simultaneously assessing the NF1 gene and a reference gene RPP30, deletions could be unambiguously distinguished from non-deletion samples. Performing the same assay for mixed samples from a DNA with a deletion and a non-deletion DNA, a highly significant linear relation was obtained between the set-up ratio of the two samples and the measured ratio of NF1/RPP30 (P<0.0001), suggesting the high potential of digital PCR in discriminating mosaic deletions. Furthermore, digital PCR detects NF1 allele loss in a tumor specimen that was not detected by loss of heterozygosity analysis using polymorphic markers due to high content of non-tumor cells. Based on the measured ratio of NF1/RPP30, the proportion of the tumor cells in this specimen could be calculated as 25%. Our results demonstrate that dual-probe digital PCR is a simple and effective method for detecting deletions and for discriminating mosaic deletions. Furthermore, this method is sensitive for assigning somatic allele loss in tumor specimen and enables determining proportion of tumor cells.</abstract><cop>England</cop><pub>Nature Publishing Group</pub><pmid>27273132</pmid><doi>10.1038/ejhg.2016.56</doi><tpages>5</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Alleles Cell Separation - methods Clonal deletion Deoxyribonucleic acid DNA DNA, Neoplasm - chemistry Gene Deletion Genetic testing Genetics Heterozygosity Humans Laboratories Loss of heterozygosity Maxillofacial surgery Mosaicism Neurofibromatosis 1 - genetics Neurofibromatosis 1 - pathology Neurofibromin 1 Neurofibromin 1 - genetics Patients Polymerase chain reaction Ratios Real-Time Polymerase Chain Reaction - methods Sensitivity and Specificity Short Report Tumor cells Tumor suppressor genes Tumors |
| title | Digital PCR for discriminating mosaic deletions and for determining proportion of tumor cells in specimen |
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