Cloning of α-β fusion gene from Clostridium perfringens and its expression
AIM： To study the cloning of α-β fusion gene from Closindium perfringens and the immunogenidty of 0-6 fusion expression. METHODS： Cloning was accomplished after PCR amplification from strains NCTC64609 and C58-1 of the protective antigen genes of α-toxin and β-toxin. The fragment of the gene was clo...
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| Veröffentlicht in: | World journal of gastroenterology : WJG 2006-02, Vol.12 (8), p.1229-1234 |
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| creator | Bai, Jia-Ning Zhang, Yan Zhao, Bao-Hua |
| description | AIM： To study the cloning of α-β fusion gene from Closindium perfringens and the immunogenidty of 0-6 fusion expression. METHODS： Cloning was accomplished after PCR amplification from strains NCTC64609 and C58-1 of the protective antigen genes of α-toxin and β-toxin. The fragment of the gene was cloned using plasmid pZCPAB. This fragment coded for the gene with the stable expression of α-β fusion gene binding. In order to verify the exact location of the α-β fusion gene, domain plasmids were constructed. The two genes were fused into expression vector pBV221. The expressed α-β fusion protein was identified by ELISA, SDS-PAGE, Western blotting and neutralization assay.
RESULTS： The protective co-toxin gene （cpa906） and the β-toxin gene （cpb930） were obtained. The recombinant plasmid pZCPAB carrying α-β fusion gene was constructed and transformed into BL21（DE3）. The recombinant strain BL21（DE3）（pZCPAB） was obtained. After the recombinant strain BL21（DE3）（pZCPAB） was induced by 42℃, its expressed product was about 22.14% of total cellular protein at SDS-PAGE and thin-layer gel scanning analysis. Neutralization assay indicated that the antibody induced by immunization with α-βfusion protein could neutralize the toxicity of α-toxin and β-toxin.
CONCLUSION： The obtained α-toxin and β-toxin genes are correct. The recombinant strain BL21（DE3）（pZCPAB） could produce α-β fusion protein. This protein can be used for immunization and is immunogenic. The antibody induced by immunization with α-β fusion protein could neutralize the toxicity of α-toxin and β-toxin. |
| doi_str_mv | 10.3748/wjg.v12.i8.1229 |
| format | Article |
| fullrecord | <record><control><sourceid>wanfang_jour_pubme</sourceid><recordid>TN_cdi_pubmedcentral_primary_oai_pubmedcentral_nih_gov_4124434</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><cqvip_id>21533173</cqvip_id><wanfj_id>wjg200608011</wanfj_id><sourcerecordid>wjg200608011</sourcerecordid><originalsourceid>FETCH-LOGICAL-c447t-19d407af62b6eb8336a8eb48b4f77997eae5d3d0c867405565b4f96d786b11e43</originalsourceid><addsrcrecordid>eNpVkMtKAzEUhoMoWi9rdxLE7dTcJslsBCneoOBG1yEzk4ypbVKTtupj6YP4TGZo8bI6i_P9_zl8ABxjNKSCyfPXSTdcYTJ0cogJqbbAgBBcFUQytA0GGCFRVJSIPbCf0gQhQmlJdsEe5iVlUvABGI-mwTvfwWDh10fx9QntMrngYWe8gTaGGcxEWkTXuuUMzk20MePGJ6h9C90iQfM2jyb1oUOwY_U0maPNPACP11cPo9tifH9zN7ocFw1jYlHgqmVIaMtJzU0tKeVamprJmlkhqkoYbcqWtqiRXDBUlrzMm4q3QvIaY8PoAbhY986X9cy0jfGLqKdqHt1Mx3cVtFP_N949qS6sFMOEMdoXnK0LXrW32ndqEpbR55dVFkoQ4kgijDN2vsaaGFKKxv6cwEj1_ntcZf_KSdX7z4mTv5_98hvhGTjdVD4F371klarWzbN1U6MILinFgtJvIJGP6A</addsrcrecordid><sourcetype>Open Access Repository</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype></control><display><type>article</type><title>Cloning of α-β fusion gene from Clostridium perfringens and its expression</title><source>MEDLINE</source><source>PubMed Central</source><source>Alma/SFX Local Collection</source><creator>Bai, Jia-Ning ; Zhang, Yan ; Zhao, Bao-Hua</creator><creatorcontrib>Bai, Jia-Ning ; Zhang, Yan ; Zhao, Bao-Hua</creatorcontrib><description>AIM： To study the cloning of α-β fusion gene from Closindium perfringens and the immunogenidty of 0-6 fusion expression. METHODS： Cloning was accomplished after PCR amplification from strains NCTC64609 and C58-1 of the protective antigen genes of α-toxin and β-toxin. The fragment of the gene was cloned using plasmid pZCPAB. This fragment coded for the gene with the stable expression of α-β fusion gene binding. In order to verify the exact location of the α-β fusion gene, domain plasmids were constructed. The two genes were fused into expression vector pBV221. The expressed α-β fusion protein was identified by ELISA, SDS-PAGE, Western blotting and neutralization assay.
RESULTS： The protective co-toxin gene （cpa906） and the β-toxin gene （cpb930） were obtained. The recombinant plasmid pZCPAB carrying α-β fusion gene was constructed and transformed into BL21（DE3）. The recombinant strain BL21（DE3）（pZCPAB） was obtained. After the recombinant strain BL21（DE3）（pZCPAB） was induced by 42℃, its expressed product was about 22.14% of total cellular protein at SDS-PAGE and thin-layer gel scanning analysis. Neutralization assay indicated that the antibody induced by immunization with α-βfusion protein could neutralize the toxicity of α-toxin and β-toxin.
CONCLUSION： The obtained α-toxin and β-toxin genes are correct. The recombinant strain BL21（DE3）（pZCPAB） could produce α-β fusion protein. This protein can be used for immunization and is immunogenic. The antibody induced by immunization with α-β fusion protein could neutralize the toxicity of α-toxin and β-toxin.</description><identifier>ISSN: 1007-9327</identifier><identifier>EISSN: 2219-2840</identifier><identifier>DOI: 10.3748/wjg.v12.i8.1229</identifier><identifier>PMID: 16534876</identifier><language>eng</language><publisher>United States: College of Life Science, Hebei Normal University, Shijiazhuang 050016 , Hebei Province, China</publisher><subject>Animals ; Bacterial Toxins - genetics ; Bacterial Toxins - immunology ; Bacterial Toxins - toxicity ; Base Sequence ; Basic Research ; Blotting, Western ; Calcium-Binding Proteins - genetics ; Calcium-Binding Proteins - immunology ; Calcium-Binding Proteins - toxicity ; Cloning, Molecular - methods ; Clostridium perfringens - genetics ; Electrophoresis, Polyacrylamide Gel ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli - genetics ; Gene Expression ; Gene Fusion ; Genes, Bacterial ; Genetic Vectors ; Mice ; Molecular Sequence Data ; Polymerase Chain Reaction ; Recombinant Fusion Proteins - genetics ; Recombinant Fusion Proteins - physiology ; Type C Phospholipases - genetics ; Type C Phospholipases - immunology ; Type C Phospholipases - toxicity ; 基因克隆 ; 基因表达 ; 梭菌 ; 细菌感染</subject><ispartof>World journal of gastroenterology : WJG, 2006-02, Vol.12 (8), p.1229-1234</ispartof><rights>Copyright © Wanfang Data Co. Ltd. All Rights Reserved.</rights><rights>2006 Baishideng Publishing Group Co., Limited. All rights reserved. 2006</rights><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c447t-19d407af62b6eb8336a8eb48b4f77997eae5d3d0c867405565b4f96d786b11e43</citedby><cites>FETCH-LOGICAL-c447t-19d407af62b6eb8336a8eb48b4f77997eae5d3d0c867405565b4f96d786b11e43</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Uhttp://image.cqvip.com/vip1000/qk/84123X/84123X.jpg</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4124434/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4124434/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,314,724,777,781,882,27906,27907,53773,53775</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/16534876$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Bai, Jia-Ning</creatorcontrib><creatorcontrib>Zhang, Yan</creatorcontrib><creatorcontrib>Zhao, Bao-Hua</creatorcontrib><title>Cloning of α-β fusion gene from Clostridium perfringens and its expression</title><title>World journal of gastroenterology : WJG</title><addtitle>World Journal of Gastroenterology</addtitle><description>AIM： To study the cloning of α-β fusion gene from Closindium perfringens and the immunogenidty of 0-6 fusion expression. METHODS： Cloning was accomplished after PCR amplification from strains NCTC64609 and C58-1 of the protective antigen genes of α-toxin and β-toxin. The fragment of the gene was cloned using plasmid pZCPAB. This fragment coded for the gene with the stable expression of α-β fusion gene binding. In order to verify the exact location of the α-β fusion gene, domain plasmids were constructed. The two genes were fused into expression vector pBV221. The expressed α-β fusion protein was identified by ELISA, SDS-PAGE, Western blotting and neutralization assay.
RESULTS： The protective co-toxin gene （cpa906） and the β-toxin gene （cpb930） were obtained. The recombinant plasmid pZCPAB carrying α-β fusion gene was constructed and transformed into BL21（DE3）. The recombinant strain BL21（DE3）（pZCPAB） was obtained. After the recombinant strain BL21（DE3）（pZCPAB） was induced by 42℃, its expressed product was about 22.14% of total cellular protein at SDS-PAGE and thin-layer gel scanning analysis. Neutralization assay indicated that the antibody induced by immunization with α-βfusion protein could neutralize the toxicity of α-toxin and β-toxin.
CONCLUSION： The obtained α-toxin and β-toxin genes are correct. The recombinant strain BL21（DE3）（pZCPAB） could produce α-β fusion protein. This protein can be used for immunization and is immunogenic. The antibody induced by immunization with α-β fusion protein could neutralize the toxicity of α-toxin and β-toxin.</description><subject>Animals</subject><subject>Bacterial Toxins - genetics</subject><subject>Bacterial Toxins - immunology</subject><subject>Bacterial Toxins - toxicity</subject><subject>Base Sequence</subject><subject>Basic Research</subject><subject>Blotting, Western</subject><subject>Calcium-Binding Proteins - genetics</subject><subject>Calcium-Binding Proteins - immunology</subject><subject>Calcium-Binding Proteins - toxicity</subject><subject>Cloning, Molecular - methods</subject><subject>Clostridium perfringens - genetics</subject><subject>Electrophoresis, Polyacrylamide Gel</subject><subject>Enzyme-Linked Immunosorbent Assay</subject><subject>Escherichia coli - genetics</subject><subject>Gene Expression</subject><subject>Gene Fusion</subject><subject>Genes, Bacterial</subject><subject>Genetic Vectors</subject><subject>Mice</subject><subject>Molecular Sequence Data</subject><subject>Polymerase Chain Reaction</subject><subject>Recombinant Fusion Proteins - genetics</subject><subject>Recombinant Fusion Proteins - physiology</subject><subject>Type C Phospholipases - genetics</subject><subject>Type C Phospholipases - immunology</subject><subject>Type C Phospholipases - toxicity</subject><subject>基因克隆</subject><subject>基因表达</subject><subject>梭菌</subject><subject>细菌感染</subject><issn>1007-9327</issn><issn>2219-2840</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2006</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpVkMtKAzEUhoMoWi9rdxLE7dTcJslsBCneoOBG1yEzk4ypbVKTtupj6YP4TGZo8bI6i_P9_zl8ABxjNKSCyfPXSTdcYTJ0cogJqbbAgBBcFUQytA0GGCFRVJSIPbCf0gQhQmlJdsEe5iVlUvABGI-mwTvfwWDh10fx9QntMrngYWe8gTaGGcxEWkTXuuUMzk20MePGJ6h9C90iQfM2jyb1oUOwY_U0maPNPACP11cPo9tifH9zN7ocFw1jYlHgqmVIaMtJzU0tKeVamprJmlkhqkoYbcqWtqiRXDBUlrzMm4q3QvIaY8PoAbhY986X9cy0jfGLqKdqHt1Mx3cVtFP_N949qS6sFMOEMdoXnK0LXrW32ndqEpbR55dVFkoQ4kgijDN2vsaaGFKKxv6cwEj1_ntcZf_KSdX7z4mTv5_98hvhGTjdVD4F371klarWzbN1U6MILinFgtJvIJGP6A</recordid><startdate>20060228</startdate><enddate>20060228</enddate><creator>Bai, Jia-Ning</creator><creator>Zhang, Yan</creator><creator>Zhao, Bao-Hua</creator><general>College of Life Science, Hebei Normal University, Shijiazhuang 050016 , Hebei Province, China</general><general>Baishideng Publishing Group Co., Limited</general><scope>2RA</scope><scope>92L</scope><scope>CQIGP</scope><scope>W91</scope><scope>~WA</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>2B.</scope><scope>4A8</scope><scope>92I</scope><scope>93N</scope><scope>PSX</scope><scope>TCJ</scope><scope>5PM</scope></search><sort><creationdate>20060228</creationdate><title>Cloning of α-β fusion gene from Clostridium perfringens and its expression</title><author>Bai, Jia-Ning ; Zhang, Yan ; Zhao, Bao-Hua</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c447t-19d407af62b6eb8336a8eb48b4f77997eae5d3d0c867405565b4f96d786b11e43</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2006</creationdate><topic>Animals</topic><topic>Bacterial Toxins - genetics</topic><topic>Bacterial Toxins - immunology</topic><topic>Bacterial Toxins - toxicity</topic><topic>Base Sequence</topic><topic>Basic Research</topic><topic>Blotting, Western</topic><topic>Calcium-Binding Proteins - genetics</topic><topic>Calcium-Binding Proteins - immunology</topic><topic>Calcium-Binding Proteins - toxicity</topic><topic>Cloning, Molecular - methods</topic><topic>Clostridium perfringens - genetics</topic><topic>Electrophoresis, Polyacrylamide Gel</topic><topic>Enzyme-Linked Immunosorbent Assay</topic><topic>Escherichia coli - genetics</topic><topic>Gene Expression</topic><topic>Gene Fusion</topic><topic>Genes, Bacterial</topic><topic>Genetic Vectors</topic><topic>Mice</topic><topic>Molecular Sequence Data</topic><topic>Polymerase Chain Reaction</topic><topic>Recombinant Fusion Proteins - genetics</topic><topic>Recombinant Fusion Proteins - physiology</topic><topic>Type C Phospholipases - genetics</topic><topic>Type C Phospholipases - immunology</topic><topic>Type C Phospholipases - toxicity</topic><topic>基因克隆</topic><topic>基因表达</topic><topic>梭菌</topic><topic>细菌感染</topic><toplevel>online_resources</toplevel><creatorcontrib>Bai, Jia-Ning</creatorcontrib><creatorcontrib>Zhang, Yan</creatorcontrib><creatorcontrib>Zhao, Bao-Hua</creatorcontrib><collection>中文科技期刊数据库</collection><collection>中文科技期刊数据库-CALIS站点</collection><collection>中文科技期刊数据库-7.0平台</collection><collection>中文科技期刊数据库-医药卫生</collection><collection>中文科技期刊数据库- 镜像站点</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Wanfang Data Journals - Hong Kong</collection><collection>WANFANG Data Centre</collection><collection>Wanfang Data Journals</collection><collection>万方数据期刊 - 香港版</collection><collection>China Online Journals (COJ)</collection><collection>China Online Journals (COJ)</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>World journal of gastroenterology : WJG</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Bai, Jia-Ning</au><au>Zhang, Yan</au><au>Zhao, Bao-Hua</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Cloning of α-β fusion gene from Clostridium perfringens and its expression</atitle><jtitle>World journal of gastroenterology : WJG</jtitle><addtitle>World Journal of Gastroenterology</addtitle><date>2006-02-28</date><risdate>2006</risdate><volume>12</volume><issue>8</issue><spage>1229</spage><epage>1234</epage><pages>1229-1234</pages><issn>1007-9327</issn><eissn>2219-2840</eissn><abstract>AIM： To study the cloning of α-β fusion gene from Closindium perfringens and the immunogenidty of 0-6 fusion expression. METHODS： Cloning was accomplished after PCR amplification from strains NCTC64609 and C58-1 of the protective antigen genes of α-toxin and β-toxin. The fragment of the gene was cloned using plasmid pZCPAB. This fragment coded for the gene with the stable expression of α-β fusion gene binding. In order to verify the exact location of the α-β fusion gene, domain plasmids were constructed. The two genes were fused into expression vector pBV221. The expressed α-β fusion protein was identified by ELISA, SDS-PAGE, Western blotting and neutralization assay.
RESULTS： The protective co-toxin gene （cpa906） and the β-toxin gene （cpb930） were obtained. The recombinant plasmid pZCPAB carrying α-β fusion gene was constructed and transformed into BL21（DE3）. The recombinant strain BL21（DE3）（pZCPAB） was obtained. After the recombinant strain BL21（DE3）（pZCPAB） was induced by 42℃, its expressed product was about 22.14% of total cellular protein at SDS-PAGE and thin-layer gel scanning analysis. Neutralization assay indicated that the antibody induced by immunization with α-βfusion protein could neutralize the toxicity of α-toxin and β-toxin.
CONCLUSION： The obtained α-toxin and β-toxin genes are correct. The recombinant strain BL21（DE3）（pZCPAB） could produce α-β fusion protein. This protein can be used for immunization and is immunogenic. The antibody induced by immunization with α-β fusion protein could neutralize the toxicity of α-toxin and β-toxin.</abstract><cop>United States</cop><pub>College of Life Science, Hebei Normal University, Shijiazhuang 050016 , Hebei Province, China</pub><pmid>16534876</pmid><doi>10.3748/wjg.v12.i8.1229</doi><tpages>6</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Animals Bacterial Toxins - genetics Bacterial Toxins - immunology Bacterial Toxins - toxicity Base Sequence Basic Research Blotting, Western Calcium-Binding Proteins - genetics Calcium-Binding Proteins - immunology Calcium-Binding Proteins - toxicity Cloning, Molecular - methods Clostridium perfringens - genetics Electrophoresis, Polyacrylamide Gel Enzyme-Linked Immunosorbent Assay Escherichia coli - genetics Gene Expression Gene Fusion Genes, Bacterial Genetic Vectors Mice Molecular Sequence Data Polymerase Chain Reaction Recombinant Fusion Proteins - genetics Recombinant Fusion Proteins - physiology Type C Phospholipases - genetics Type C Phospholipases - immunology Type C Phospholipases - toxicity 基因克隆 基因表达 梭菌 细菌感染 |
| title | Cloning of α-β fusion gene from Clostridium perfringens and its expression |
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