Using Next Generation Sequencing for Multiplexed Trait-Linked Markers in Wheat
With the advent of next generation sequencing (NGS) technologies, single nucleotide polymorphisms (SNPs) have become the major type of marker for genotyping in many crops. However, the availability of SNP markers for important traits of bread wheat (Triticum aestivum L.) that can be effectively used...
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| creator | Bernardo, Amy Wang, Shan St Amand, Paul Bai, Guihua |
| description | With the advent of next generation sequencing (NGS) technologies, single nucleotide polymorphisms (SNPs) have become the major type of marker for genotyping in many crops. However, the availability of SNP markers for important traits of bread wheat (Triticum aestivum L.) that can be effectively used in marker-assisted selection (MAS) is still limited and SNP assays for MAS are usually uniplex. A shift from uniplex to multiplex assays will allow the simultaneous analysis of multiple markers and increase MAS efficiency. We designed 33 locus-specific markers from SNP or indel-based marker sequences that linked to 20 different quantitative trait loci (QTL) or genes of agronomic importance in wheat and analyzed the amplicon sequences using an Ion Torrent Proton Sequencer and a custom allele detection pipeline to determine the genotypes of 24 selected germplasm accessions. Among the 33 markers, 27 were successfully multiplexed and 23 had 100% SNP call rates. Results from analysis of "kompetitive allele-specific PCR" (KASP) and sequence tagged site (STS) markers developed from the same loci fully verified the genotype calls of 23 markers. The NGS-based multiplexed assay developed in this study is suitable for rapid and high-throughput screening of SNPs and some indel-based markers in wheat. |
| doi_str_mv | 10.1371/journal.pone.0143890 |
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However, the availability of SNP markers for important traits of bread wheat (Triticum aestivum L.) that can be effectively used in marker-assisted selection (MAS) is still limited and SNP assays for MAS are usually uniplex. A shift from uniplex to multiplex assays will allow the simultaneous analysis of multiple markers and increase MAS efficiency. We designed 33 locus-specific markers from SNP or indel-based marker sequences that linked to 20 different quantitative trait loci (QTL) or genes of agronomic importance in wheat and analyzed the amplicon sequences using an Ion Torrent Proton Sequencer and a custom allele detection pipeline to determine the genotypes of 24 selected germplasm accessions. Among the 33 markers, 27 were successfully multiplexed and 23 had 100% SNP call rates. Results from analysis of "kompetitive allele-specific PCR" (KASP) and sequence tagged site (STS) markers developed from the same loci fully verified the genotype calls of 23 markers. The NGS-based multiplexed assay developed in this study is suitable for rapid and high-throughput screening of SNPs and some indel-based markers in wheat.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0143890</identifier><identifier>PMID: 26625271</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Agriculture ; Agronomy ; Alleles ; Analysis ; Assaying ; Bread ; Chromosome Mapping - methods ; Chromosomes ; Crops, Agricultural - genetics ; Deoxyribonucleic acid ; DNA ; Gene sequencing ; Genes ; Genes, Plant - genetics ; Genetic Linkage - genetics ; Genetic Markers - genetics ; Genetics ; Genomes ; Genomics ; Genotype ; Genotypes ; Genotyping ; Germplasm ; High-Throughput Nucleotide Sequencing - methods ; High-throughput screening ; Marker-assisted selection ; Markers ; Multiplexing ; Oryza ; Phenotype ; Phenotypes ; Polymorphism, Single Nucleotide - genetics ; Quantitative genetics ; Quantitative trait loci ; Quantitative Trait Loci - genetics ; Rice ; Sequence Tagged Sites ; Single nucleotide polymorphisms ; Single-nucleotide polymorphism ; Triticum - genetics ; Triticum aestivum ; Wheat</subject><ispartof>PloS one, 2015-12, Vol.10 (12), p.e0143890-e0143890</ispartof><rights>COPYRIGHT 2015 Public Library of Science</rights><rights>This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication: https://creativecommons.org/publicdomain/zero/1.0/ (the “License”) Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c758t-65b65efe07628372c29f01eb911ea6d4ff891f164d1946953fa6f304c5e7b7203</citedby><cites>FETCH-LOGICAL-c758t-65b65efe07628372c29f01eb911ea6d4ff891f164d1946953fa6f304c5e7b7203</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/PMC4666610/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4666610/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,860,881,2095,2914,23846,27903,27904,53769,53771,79346,79347</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/26625271$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Fang, David D</contributor><creatorcontrib>Bernardo, Amy</creatorcontrib><creatorcontrib>Wang, Shan</creatorcontrib><creatorcontrib>St Amand, Paul</creatorcontrib><creatorcontrib>Bai, Guihua</creatorcontrib><title>Using Next Generation Sequencing for Multiplexed Trait-Linked Markers in Wheat</title><title>PloS one</title><addtitle>PLoS One</addtitle><description>With the advent of next generation sequencing (NGS) technologies, single nucleotide polymorphisms (SNPs) have become the major type of marker for genotyping in many crops. However, the availability of SNP markers for important traits of bread wheat (Triticum aestivum L.) that can be effectively used in marker-assisted selection (MAS) is still limited and SNP assays for MAS are usually uniplex. A shift from uniplex to multiplex assays will allow the simultaneous analysis of multiple markers and increase MAS efficiency. We designed 33 locus-specific markers from SNP or indel-based marker sequences that linked to 20 different quantitative trait loci (QTL) or genes of agronomic importance in wheat and analyzed the amplicon sequences using an Ion Torrent Proton Sequencer and a custom allele detection pipeline to determine the genotypes of 24 selected germplasm accessions. Among the 33 markers, 27 were successfully multiplexed and 23 had 100% SNP call rates. Results from analysis of "kompetitive allele-specific PCR" (KASP) and sequence tagged site (STS) markers developed from the same loci fully verified the genotype calls of 23 markers. The NGS-based multiplexed assay developed in this study is suitable for rapid and high-throughput screening of SNPs and some indel-based markers in wheat.</description><subject>Agriculture</subject><subject>Agronomy</subject><subject>Alleles</subject><subject>Analysis</subject><subject>Assaying</subject><subject>Bread</subject><subject>Chromosome Mapping - methods</subject><subject>Chromosomes</subject><subject>Crops, Agricultural - genetics</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>Gene sequencing</subject><subject>Genes</subject><subject>Genes, Plant - genetics</subject><subject>Genetic Linkage - genetics</subject><subject>Genetic Markers - genetics</subject><subject>Genetics</subject><subject>Genomes</subject><subject>Genomics</subject><subject>Genotype</subject><subject>Genotypes</subject><subject>Genotyping</subject><subject>Germplasm</subject><subject>High-Throughput Nucleotide Sequencing - methods</subject><subject>High-throughput screening</subject><subject>Marker-assisted selection</subject><subject>Markers</subject><subject>Multiplexing</subject><subject>Oryza</subject><subject>Phenotype</subject><subject>Phenotypes</subject><subject>Polymorphism, Single Nucleotide - genetics</subject><subject>Quantitative genetics</subject><subject>Quantitative trait loci</subject><subject>Quantitative Trait Loci - genetics</subject><subject>Rice</subject><subject>Sequence Tagged Sites</subject><subject>Single nucleotide polymorphisms</subject><subject>Single-nucleotide polymorphism</subject><subject>Triticum - genetics</subject><subject>Triticum aestivum</subject><subject>Wheat</subject><issn>1932-6203</issn><issn>1932-6203</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</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><sourceid>DOA</sourceid><recordid>eNqNkl1v0zAUhiMEYmPwDxBEQkJw0RJ_xE5uJk0TjErdJrENLi3XOW7duXaxEzT-PQ7Npgbtgvgi_njO63OO3yx7jYopIhx9WvsuOGmnW-9gWiBKqrp4kh2imuAJwwV5ujc_yF7EuC6KklSMPc8OMGO4xBwdZhc30bhlfgF3bX4GDoJsjXf5FfzswKn-SPuQn3e2NVsLd9Dk10GadjI37jYtzmW4hRBz4_IfK5Dty-yZljbCq-F_lN18-Xx9-nUyvzybnZ7MJ4qXVTth5YKVoKHgDFeEY4VrXSBY1AiBZA3VuqqRRow2qKasLomWTJOCqhL4gqeCjrK3O92t9VEMrYgCcVJRznfEbEc0Xq7FNpiNDL-Fl0b83fBhKWRojbIgZAW1VJQjpigFiirMJFuknEqsFGvqpHU83NYtNtAocG2QdiQ6PnFmJZb-l6AsfahP5sMgEHxqbGzFxkQF1koHvtvlXWFUoiqh7_5BH69uoJYyFWCc9ule1YuKE0oY54wUfd7TR6g0GtgYlXyjTdofBXwcBSSmTc5Yyi5GMbv69v_s5fcx-36PTUax7Sp62_Vei2OQ7kAVfIwB9EOTUSF62993Q_S2F4PtU9ib_Qd6CLr3OfkDLwH7fw</recordid><startdate>20151201</startdate><enddate>20151201</enddate><creator>Bernardo, Amy</creator><creator>Wang, Shan</creator><creator>St Amand, Paul</creator><creator>Bai, Guihua</creator><general>Public Library of Science</general><general>Public Library of Science (PLoS)</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>IOV</scope><scope>ISR</scope><scope>3V.</scope><scope>7QG</scope><scope>7QL</scope><scope>7QO</scope><scope>7RV</scope><scope>7SN</scope><scope>7SS</scope><scope>7T5</scope><scope>7TG</scope><scope>7TM</scope><scope>7U9</scope><scope>7X2</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8C1</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABJCF</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>ARAPS</scope><scope>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</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>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>KB.</scope><scope>KB0</scope><scope>KL.</scope><scope>L6V</scope><scope>LK8</scope><scope>M0K</scope><scope>M0S</scope><scope>M1P</scope><scope>M7N</scope><scope>M7P</scope><scope>M7S</scope><scope>NAPCQ</scope><scope>P5Z</scope><scope>P62</scope><scope>P64</scope><scope>PATMY</scope><scope>PDBOC</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>PTHSS</scope><scope>PYCSY</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope><scope>DOA</scope></search><sort><creationdate>20151201</creationdate><title>Using Next Generation Sequencing for Multiplexed Trait-Linked Markers in Wheat</title><author>Bernardo, Amy ; Wang, Shan ; St Amand, Paul ; Bai, Guihua</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c758t-65b65efe07628372c29f01eb911ea6d4ff891f164d1946953fa6f304c5e7b7203</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Agriculture</topic><topic>Agronomy</topic><topic>Alleles</topic><topic>Analysis</topic><topic>Assaying</topic><topic>Bread</topic><topic>Chromosome Mapping - methods</topic><topic>Chromosomes</topic><topic>Crops, Agricultural - genetics</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>Gene sequencing</topic><topic>Genes</topic><topic>Genes, Plant - genetics</topic><topic>Genetic Linkage - genetics</topic><topic>Genetic Markers - genetics</topic><topic>Genetics</topic><topic>Genomes</topic><topic>Genomics</topic><topic>Genotype</topic><topic>Genotypes</topic><topic>Genotyping</topic><topic>Germplasm</topic><topic>High-Throughput Nucleotide Sequencing - methods</topic><topic>High-throughput screening</topic><topic>Marker-assisted selection</topic><topic>Markers</topic><topic>Multiplexing</topic><topic>Oryza</topic><topic>Phenotype</topic><topic>Phenotypes</topic><topic>Polymorphism, Single Nucleotide - genetics</topic><topic>Quantitative genetics</topic><topic>Quantitative trait loci</topic><topic>Quantitative Trait Loci - genetics</topic><topic>Rice</topic><topic>Sequence Tagged Sites</topic><topic>Single nucleotide polymorphisms</topic><topic>Single-nucleotide polymorphism</topic><topic>Triticum - genetics</topic><topic>Triticum aestivum</topic><topic>Wheat</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Bernardo, Amy</creatorcontrib><creatorcontrib>Wang, Shan</creatorcontrib><creatorcontrib>St Amand, Paul</creatorcontrib><creatorcontrib>Bai, Guihua</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Gale In Context: Opposing Viewpoints</collection><collection>Gale In Context: Science</collection><collection>ProQuest Central (Corporate)</collection><collection>Animal Behavior Abstracts</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Biotechnology Research Abstracts</collection><collection>Proquest Nursing & Allied Health Source</collection><collection>Ecology Abstracts</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Immunology Abstracts</collection><collection>Meteorological & Geoastrophysical Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Agricultural Science Collection</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical 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>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>Materials Science & Engineering Collection</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest One Sustainability</collection><collection>ProQuest Central UK/Ireland</collection><collection>Advanced Technologies & Aerospace Collection</collection><collection>Agricultural & Environmental Science Collection</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Technology Collection (ProQuest)</collection><collection>Natural Science Collection (ProQuest)</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>AIDS and Cancer Research Abstracts</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>Meteorological & Geoastrophysical Abstracts - 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Academic</collection><collection>PubMed Central (Full Participant titles)</collection><collection>DOAJ Directory of Open Access Journals</collection><jtitle>PloS one</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Bernardo, Amy</au><au>Wang, Shan</au><au>St Amand, Paul</au><au>Bai, Guihua</au><au>Fang, David D</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Using Next Generation Sequencing for Multiplexed Trait-Linked Markers in Wheat</atitle><jtitle>PloS one</jtitle><addtitle>PLoS One</addtitle><date>2015-12-01</date><risdate>2015</risdate><volume>10</volume><issue>12</issue><spage>e0143890</spage><epage>e0143890</epage><pages>e0143890-e0143890</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>With the advent of next generation sequencing (NGS) technologies, single nucleotide polymorphisms (SNPs) have become the major type of marker for genotyping in many crops. However, the availability of SNP markers for important traits of bread wheat (Triticum aestivum L.) that can be effectively used in marker-assisted selection (MAS) is still limited and SNP assays for MAS are usually uniplex. A shift from uniplex to multiplex assays will allow the simultaneous analysis of multiple markers and increase MAS efficiency. We designed 33 locus-specific markers from SNP or indel-based marker sequences that linked to 20 different quantitative trait loci (QTL) or genes of agronomic importance in wheat and analyzed the amplicon sequences using an Ion Torrent Proton Sequencer and a custom allele detection pipeline to determine the genotypes of 24 selected germplasm accessions. Among the 33 markers, 27 were successfully multiplexed and 23 had 100% SNP call rates. Results from analysis of "kompetitive allele-specific PCR" (KASP) and sequence tagged site (STS) markers developed from the same loci fully verified the genotype calls of 23 markers. The NGS-based multiplexed assay developed in this study is suitable for rapid and high-throughput screening of SNPs and some indel-based markers in wheat.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>26625271</pmid><doi>10.1371/journal.pone.0143890</doi><oa>free_for_read</oa></addata></record> |
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| subjects | Agriculture Agronomy Alleles Analysis Assaying Bread Chromosome Mapping - methods Chromosomes Crops, Agricultural - genetics Deoxyribonucleic acid DNA Gene sequencing Genes Genes, Plant - genetics Genetic Linkage - genetics Genetic Markers - genetics Genetics Genomes Genomics Genotype Genotypes Genotyping Germplasm High-Throughput Nucleotide Sequencing - methods High-throughput screening Marker-assisted selection Markers Multiplexing Oryza Phenotype Phenotypes Polymorphism, Single Nucleotide - genetics Quantitative genetics Quantitative trait loci Quantitative Trait Loci - genetics Rice Sequence Tagged Sites Single nucleotide polymorphisms Single-nucleotide polymorphism Triticum - genetics Triticum aestivum Wheat |
| title | Using Next Generation Sequencing for Multiplexed Trait-Linked Markers in Wheat |
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