Highly Sensitive Pyrosequencing System with Polymer-Supported Enzymes for High-Throughput DNA Analysis
A highly sensitive massively parallel pyrosequencing system employing a gel matrix to immobilize enzymes at high density in microreaction chambers is demonstrated. Reducing the size of microreaction chambers in a DNA analyzer is important to achieve a high throughput utilizing a commercially availab...
Gespeichert in:
| Veröffentlicht in: | Analytical chemistry (Washington) 2011-10, Vol.83 (19), p.7560-7565 |
|---|---|
| Hauptverfasser: | , , , , , |
| Format: | Artikel |
| Sprache: | eng |
| Schlagworte: | |
| Online-Zugang: | Volltext |
| Tags: |
Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
|
| container_end_page | 7565 |
|---|---|
| container_issue | 19 |
| container_start_page | 7560 |
| container_title | Analytical chemistry (Washington) |
| container_volume | 83 |
| creator | Shirai, Masataka Goto, Mari Suzuki, Shigeya Kono, Kenji Kajiyama, Tomoharu Kambara, Hideki |
| description | A highly sensitive massively parallel pyrosequencing system employing a gel matrix to immobilize enzymes at high density in microreaction chambers is demonstrated. Reducing the size of microreaction chambers in a DNA analyzer is important to achieve a high throughput utilizing a commercially available detection device or camera. A high-performance system can be attained by detecting signals from one reaction chamber with one photopixel of around several micrometers by utilizing a 1:1 image magnification. However, the use of small beads immobilizing DNA has a disadvantage in detecting luminescence because only small amounts of DNA can be immobilized on the bead surfaces for sequencing. As luminescence intensity could be enhanced by increasing the luciferase density in the chambers, we overcame this difficulty by using a gel matrix to immobilize luciferase at a high concentration in the microreaction chambers. Luminescence 1 order of magnitude higher could be observed with the new method compared to the conventional method. Consequently, the chamber size and bead size immobilizing DNA could be reduced to as small as 6.5 and 4 μm, respectively. This can be successfully applied to achieving small, inexpensive, pyrosequencing systems with high throughput. |
| doi_str_mv | 10.1021/ac201357t |
| format | Article |
| fullrecord | <record><control><sourceid>proquest_cross</sourceid><recordid>TN_cdi_proquest_miscellaneous_895857422</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>895857422</sourcerecordid><originalsourceid>FETCH-LOGICAL-a371t-25aca79a384ce313e37f4a6956d01b2ac478768cceb46d0da60b7c8f79588d4f3</originalsourceid><addsrcrecordid>eNpl0MtKxDAUBuAgio6XhS8gQRBxUT1Jc-tyGK8gKoyuSyZNZyq9mbRKfXozODqgq8Dh4z8nP0KHBM4JUHKhDQUSc9ltoBHhFCKhFN1EIwCIIyoBdtCu968AhAAR22iHEsUZcBih_LaYL8oBT23ti654t_hpcI23b72tTVHP8XTwna3wR9Et8FNTDpV10bRv28Z1NsNX9WeYeJw3Di-ToueFa_r5ou07fPkwxuNal4Mv_D7aynXp7cHq3UMv11fPk9vo_vHmbjK-j3QsSRdRro2WiY4VMzYmsY1lzrRIuMiAzKg2TCoplDF2xsIo0wJm0qhcJlypjOXxHjr9zm1dE77gu7QqvLFlqWvb9D5VAXLJKA3y-I98bXoXzl2iRDAGIgno7BuZ0Il3Nk9bV1TaDSmBdFl9-lt9sEerwH5W2exX_nQdwMkKaG90mTsdGvZrx7hIpKBrp41fH_V_4Re5U5es</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>899644069</pqid></control><display><type>article</type><title>Highly Sensitive Pyrosequencing System with Polymer-Supported Enzymes for High-Throughput DNA Analysis</title><source>MEDLINE</source><source>ACS Publications</source><creator>Shirai, Masataka ; Goto, Mari ; Suzuki, Shigeya ; Kono, Kenji ; Kajiyama, Tomoharu ; Kambara, Hideki</creator><creatorcontrib>Shirai, Masataka ; Goto, Mari ; Suzuki, Shigeya ; Kono, Kenji ; Kajiyama, Tomoharu ; Kambara, Hideki</creatorcontrib><description>A highly sensitive massively parallel pyrosequencing system employing a gel matrix to immobilize enzymes at high density in microreaction chambers is demonstrated. Reducing the size of microreaction chambers in a DNA analyzer is important to achieve a high throughput utilizing a commercially available detection device or camera. A high-performance system can be attained by detecting signals from one reaction chamber with one photopixel of around several micrometers by utilizing a 1:1 image magnification. However, the use of small beads immobilizing DNA has a disadvantage in detecting luminescence because only small amounts of DNA can be immobilized on the bead surfaces for sequencing. As luminescence intensity could be enhanced by increasing the luciferase density in the chambers, we overcame this difficulty by using a gel matrix to immobilize luciferase at a high concentration in the microreaction chambers. Luminescence 1 order of magnitude higher could be observed with the new method compared to the conventional method. Consequently, the chamber size and bead size immobilizing DNA could be reduced to as small as 6.5 and 4 μm, respectively. This can be successfully applied to achieving small, inexpensive, pyrosequencing systems with high throughput.</description><identifier>ISSN: 0003-2700</identifier><identifier>EISSN: 1520-6882</identifier><identifier>DOI: 10.1021/ac201357t</identifier><identifier>PMID: 21854050</identifier><identifier>CODEN: ANCHAM</identifier><language>eng</language><publisher>Washington, DC: American Chemical Society</publisher><subject>Analytical chemistry ; Analyzers ; Chemistry ; Deoxyribonucleic acid ; DNA ; DNA - analysis ; DNA - genetics ; Enzymes ; Enzymes, Immobilized - chemistry ; Enzymes, Immobilized - metabolism ; Exact sciences and technology ; High-Throughput Nucleotide Sequencing - methods ; Luciferases - chemistry ; Luciferases - metabolism ; Luminescence ; Polymers - chemistry ; Spectrometric and optical methods ; Sulfate Adenylyltransferase - chemistry ; Sulfate Adenylyltransferase - metabolism</subject><ispartof>Analytical chemistry (Washington), 2011-10, Vol.83 (19), p.7560-7565</ispartof><rights>Copyright © 2011 American Chemical Society</rights><rights>2015 INIST-CNRS</rights><rights>Copyright American Chemical Society Oct 1, 2011</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a371t-25aca79a384ce313e37f4a6956d01b2ac478768cceb46d0da60b7c8f79588d4f3</citedby><cites>FETCH-LOGICAL-a371t-25aca79a384ce313e37f4a6956d01b2ac478768cceb46d0da60b7c8f79588d4f3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://pubs.acs.org/doi/pdf/10.1021/ac201357t$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/ac201357t$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,780,784,2765,27076,27924,27925,56738,56788</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=24569762$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/21854050$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Shirai, Masataka</creatorcontrib><creatorcontrib>Goto, Mari</creatorcontrib><creatorcontrib>Suzuki, Shigeya</creatorcontrib><creatorcontrib>Kono, Kenji</creatorcontrib><creatorcontrib>Kajiyama, Tomoharu</creatorcontrib><creatorcontrib>Kambara, Hideki</creatorcontrib><title>Highly Sensitive Pyrosequencing System with Polymer-Supported Enzymes for High-Throughput DNA Analysis</title><title>Analytical chemistry (Washington)</title><addtitle>Anal. Chem</addtitle><description>A highly sensitive massively parallel pyrosequencing system employing a gel matrix to immobilize enzymes at high density in microreaction chambers is demonstrated. Reducing the size of microreaction chambers in a DNA analyzer is important to achieve a high throughput utilizing a commercially available detection device or camera. A high-performance system can be attained by detecting signals from one reaction chamber with one photopixel of around several micrometers by utilizing a 1:1 image magnification. However, the use of small beads immobilizing DNA has a disadvantage in detecting luminescence because only small amounts of DNA can be immobilized on the bead surfaces for sequencing. As luminescence intensity could be enhanced by increasing the luciferase density in the chambers, we overcame this difficulty by using a gel matrix to immobilize luciferase at a high concentration in the microreaction chambers. Luminescence 1 order of magnitude higher could be observed with the new method compared to the conventional method. Consequently, the chamber size and bead size immobilizing DNA could be reduced to as small as 6.5 and 4 μm, respectively. This can be successfully applied to achieving small, inexpensive, pyrosequencing systems with high throughput.</description><subject>Analytical chemistry</subject><subject>Analyzers</subject><subject>Chemistry</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>DNA - analysis</subject><subject>DNA - genetics</subject><subject>Enzymes</subject><subject>Enzymes, Immobilized - chemistry</subject><subject>Enzymes, Immobilized - metabolism</subject><subject>Exact sciences and technology</subject><subject>High-Throughput Nucleotide Sequencing - methods</subject><subject>Luciferases - chemistry</subject><subject>Luciferases - metabolism</subject><subject>Luminescence</subject><subject>Polymers - chemistry</subject><subject>Spectrometric and optical methods</subject><subject>Sulfate Adenylyltransferase - chemistry</subject><subject>Sulfate Adenylyltransferase - metabolism</subject><issn>0003-2700</issn><issn>1520-6882</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2011</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpl0MtKxDAUBuAgio6XhS8gQRBxUT1Jc-tyGK8gKoyuSyZNZyq9mbRKfXozODqgq8Dh4z8nP0KHBM4JUHKhDQUSc9ltoBHhFCKhFN1EIwCIIyoBdtCu968AhAAR22iHEsUZcBih_LaYL8oBT23ti654t_hpcI23b72tTVHP8XTwna3wR9Et8FNTDpV10bRv28Z1NsNX9WeYeJw3Di-ToueFa_r5ou07fPkwxuNal4Mv_D7aynXp7cHq3UMv11fPk9vo_vHmbjK-j3QsSRdRro2WiY4VMzYmsY1lzrRIuMiAzKg2TCoplDF2xsIo0wJm0qhcJlypjOXxHjr9zm1dE77gu7QqvLFlqWvb9D5VAXLJKA3y-I98bXoXzl2iRDAGIgno7BuZ0Il3Nk9bV1TaDSmBdFl9-lt9sEerwH5W2exX_nQdwMkKaG90mTsdGvZrx7hIpKBrp41fH_V_4Re5U5es</recordid><startdate>20111001</startdate><enddate>20111001</enddate><creator>Shirai, Masataka</creator><creator>Goto, Mari</creator><creator>Suzuki, Shigeya</creator><creator>Kono, Kenji</creator><creator>Kajiyama, Tomoharu</creator><creator>Kambara, Hideki</creator><general>American Chemical Society</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>7QF</scope><scope>7QO</scope><scope>7QQ</scope><scope>7SC</scope><scope>7SE</scope><scope>7SP</scope><scope>7SR</scope><scope>7TA</scope><scope>7TB</scope><scope>7TM</scope><scope>7U5</scope><scope>7U7</scope><scope>7U9</scope><scope>8BQ</scope><scope>8FD</scope><scope>C1K</scope><scope>F28</scope><scope>FR3</scope><scope>H8D</scope><scope>H8G</scope><scope>H94</scope><scope>JG9</scope><scope>JQ2</scope><scope>KR7</scope><scope>L7M</scope><scope>L~C</scope><scope>L~D</scope><scope>P64</scope><scope>7X8</scope></search><sort><creationdate>20111001</creationdate><title>Highly Sensitive Pyrosequencing System with Polymer-Supported Enzymes for High-Throughput DNA Analysis</title><author>Shirai, Masataka ; Goto, Mari ; Suzuki, Shigeya ; Kono, Kenji ; Kajiyama, Tomoharu ; Kambara, Hideki</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a371t-25aca79a384ce313e37f4a6956d01b2ac478768cceb46d0da60b7c8f79588d4f3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2011</creationdate><topic>Analytical chemistry</topic><topic>Analyzers</topic><topic>Chemistry</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>DNA - analysis</topic><topic>DNA - genetics</topic><topic>Enzymes</topic><topic>Enzymes, Immobilized - chemistry</topic><topic>Enzymes, Immobilized - metabolism</topic><topic>Exact sciences and technology</topic><topic>High-Throughput Nucleotide Sequencing - methods</topic><topic>Luciferases - chemistry</topic><topic>Luciferases - metabolism</topic><topic>Luminescence</topic><topic>Polymers - chemistry</topic><topic>Spectrometric and optical methods</topic><topic>Sulfate Adenylyltransferase - chemistry</topic><topic>Sulfate Adenylyltransferase - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Shirai, Masataka</creatorcontrib><creatorcontrib>Goto, Mari</creatorcontrib><creatorcontrib>Suzuki, Shigeya</creatorcontrib><creatorcontrib>Kono, Kenji</creatorcontrib><creatorcontrib>Kajiyama, Tomoharu</creatorcontrib><creatorcontrib>Kambara, Hideki</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>Aluminium Industry Abstracts</collection><collection>Biotechnology Research Abstracts</collection><collection>Ceramic Abstracts</collection><collection>Computer and Information Systems Abstracts</collection><collection>Corrosion Abstracts</collection><collection>Electronics & Communications Abstracts</collection><collection>Engineered Materials Abstracts</collection><collection>Materials Business File</collection><collection>Mechanical & Transportation Engineering Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Solid State and Superconductivity Abstracts</collection><collection>Toxicology Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>METADEX</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ANTE: Abstracts in New Technology & Engineering</collection><collection>Engineering Research Database</collection><collection>Aerospace Database</collection><collection>Copper Technical Reference Library</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Materials Research Database</collection><collection>ProQuest Computer Science Collection</collection><collection>Civil Engineering Abstracts</collection><collection>Advanced Technologies Database with Aerospace</collection><collection>Computer and Information Systems Abstracts Academic</collection><collection>Computer and Information Systems Abstracts Professional</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Analytical chemistry (Washington)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Shirai, Masataka</au><au>Goto, Mari</au><au>Suzuki, Shigeya</au><au>Kono, Kenji</au><au>Kajiyama, Tomoharu</au><au>Kambara, Hideki</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Highly Sensitive Pyrosequencing System with Polymer-Supported Enzymes for High-Throughput DNA Analysis</atitle><jtitle>Analytical chemistry (Washington)</jtitle><addtitle>Anal. Chem</addtitle><date>2011-10-01</date><risdate>2011</risdate><volume>83</volume><issue>19</issue><spage>7560</spage><epage>7565</epage><pages>7560-7565</pages><issn>0003-2700</issn><eissn>1520-6882</eissn><coden>ANCHAM</coden><abstract>A highly sensitive massively parallel pyrosequencing system employing a gel matrix to immobilize enzymes at high density in microreaction chambers is demonstrated. Reducing the size of microreaction chambers in a DNA analyzer is important to achieve a high throughput utilizing a commercially available detection device or camera. A high-performance system can be attained by detecting signals from one reaction chamber with one photopixel of around several micrometers by utilizing a 1:1 image magnification. However, the use of small beads immobilizing DNA has a disadvantage in detecting luminescence because only small amounts of DNA can be immobilized on the bead surfaces for sequencing. As luminescence intensity could be enhanced by increasing the luciferase density in the chambers, we overcame this difficulty by using a gel matrix to immobilize luciferase at a high concentration in the microreaction chambers. Luminescence 1 order of magnitude higher could be observed with the new method compared to the conventional method. Consequently, the chamber size and bead size immobilizing DNA could be reduced to as small as 6.5 and 4 μm, respectively. This can be successfully applied to achieving small, inexpensive, pyrosequencing systems with high throughput.</abstract><cop>Washington, DC</cop><pub>American Chemical Society</pub><pmid>21854050</pmid><doi>10.1021/ac201357t</doi><tpages>6</tpages></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0003-2700 |
| ispartof | Analytical chemistry (Washington), 2011-10, Vol.83 (19), p.7560-7565 |
| issn | 0003-2700 1520-6882 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_895857422 |
| source | MEDLINE; ACS Publications |
| subjects | Analytical chemistry Analyzers Chemistry Deoxyribonucleic acid DNA DNA - analysis DNA - genetics Enzymes Enzymes, Immobilized - chemistry Enzymes, Immobilized - metabolism Exact sciences and technology High-Throughput Nucleotide Sequencing - methods Luciferases - chemistry Luciferases - metabolism Luminescence Polymers - chemistry Spectrometric and optical methods Sulfate Adenylyltransferase - chemistry Sulfate Adenylyltransferase - metabolism |
| title | Highly Sensitive Pyrosequencing System with Polymer-Supported Enzymes for High-Throughput DNA Analysis |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-06T19%3A08%3A47IST&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=Highly%20Sensitive%20Pyrosequencing%20System%20with%20Polymer-Supported%20Enzymes%20for%20High-Throughput%20DNA%20Analysis&rft.jtitle=Analytical%20chemistry%20(Washington)&rft.au=Shirai,%20Masataka&rft.date=2011-10-01&rft.volume=83&rft.issue=19&rft.spage=7560&rft.epage=7565&rft.pages=7560-7565&rft.issn=0003-2700&rft.eissn=1520-6882&rft.coden=ANCHAM&rft_id=info:doi/10.1021/ac201357t&rft_dat=%3Cproquest_cross%3E895857422%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=899644069&rft_id=info:pmid/21854050&rfr_iscdi=true |