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Article: Cloning, purification, and antigenic characterization of three recombinant fragments derived from SARS-CoV S1 domain

TitleCloning, purification, and antigenic characterization of three recombinant fragments derived from SARS-CoV S1 domain
Authors
Issue Date2005
Citation
Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi = Zhonghua Shiyan He Linchuang Bingduxue Zazhi = Chinese Journal Of Experimental And Clinical Virology, 2005, v. 19 n. 3, p. 275-278 How to Cite?
AbstractOBJECTIVE: The present study aimed to clone and express three fragments of genomic RNA derived from SARS associated coronavirus (SARS-CoV) S1 domain and to study its immunogenicity. METHODS: The S1 domain gene was amplified by PCR with specific primers and was inserted into the prokaryotic expression vector pQE-30. Three fragments (40-751, 746-1344 and 746-2001 bp) derived from S1 domain produced after the recombinant plasmid (pQE-30/S1) was digested by restriction endonucleases. The three fragments were cloned into pQE-30 and expressed in M15 strains of Escherichia coli. The expression products, designated S1a, S1b and S1c respectively, were purified by Ni affinity chromatography. The immunogenicity was analyzed by Western Blot and ELISA using serologically confirmed sera from SARS patients and the sera from healthy donors was used as control at the same assay. RESULTS: Three recombinant plasmids (pQE-30/S1a, pQE-30/S1b, pQE-30/S1c) were constructed.Fusion proteins with relative molecular mass of 26,700, 22,500 and 46,000 dalton were successfully expressed with amounts of 35%, 35% and 30% of total cell protein and purified by Ni affinity chromatography, respectively. Western Blot and ELISA analysis showed that the S1c protein could be specifically recognized by the sera from SARS patients. CONCLUSION: The recombinant S1c protein was a good immunogen and has the potential to be used as a vaccine against SARS-CoV infection.
Persistent Identifierhttp://hdl.handle.net/10722/157536
ISSN
2015 SCImago Journal Rankings: 0.140

 

DC FieldValueLanguage
dc.contributor.authorMei, YBen_US
dc.contributor.authorLiao, ZYen_US
dc.contributor.authorWang, YDen_US
dc.contributor.authorZhang, LYen_US
dc.contributor.authorXu, Hen_US
dc.contributor.authorYuen, KYen_US
dc.contributor.authorChe, XYen_US
dc.date.accessioned2012-08-08T08:51:03Z-
dc.date.available2012-08-08T08:51:03Z-
dc.date.issued2005en_US
dc.identifier.citationZhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi = Zhonghua Shiyan He Linchuang Bingduxue Zazhi = Chinese Journal Of Experimental And Clinical Virology, 2005, v. 19 n. 3, p. 275-278en_US
dc.identifier.issn1003-9279en_US
dc.identifier.urihttp://hdl.handle.net/10722/157536-
dc.description.abstractOBJECTIVE: The present study aimed to clone and express three fragments of genomic RNA derived from SARS associated coronavirus (SARS-CoV) S1 domain and to study its immunogenicity. METHODS: The S1 domain gene was amplified by PCR with specific primers and was inserted into the prokaryotic expression vector pQE-30. Three fragments (40-751, 746-1344 and 746-2001 bp) derived from S1 domain produced after the recombinant plasmid (pQE-30/S1) was digested by restriction endonucleases. The three fragments were cloned into pQE-30 and expressed in M15 strains of Escherichia coli. The expression products, designated S1a, S1b and S1c respectively, were purified by Ni affinity chromatography. The immunogenicity was analyzed by Western Blot and ELISA using serologically confirmed sera from SARS patients and the sera from healthy donors was used as control at the same assay. RESULTS: Three recombinant plasmids (pQE-30/S1a, pQE-30/S1b, pQE-30/S1c) were constructed.Fusion proteins with relative molecular mass of 26,700, 22,500 and 46,000 dalton were successfully expressed with amounts of 35%, 35% and 30% of total cell protein and purified by Ni affinity chromatography, respectively. Western Blot and ELISA analysis showed that the S1c protein could be specifically recognized by the sera from SARS patients. CONCLUSION: The recombinant S1c protein was a good immunogen and has the potential to be used as a vaccine against SARS-CoV infection.en_US
dc.languageengen_US
dc.relation.ispartofZhonghua shi yan he lin chuang bing du xue za zhi = Zhonghua shiyan he linchuang bingduxue zazhi = Chinese journal of experimental and clinical virologyen_US
dc.subject.meshAntibodies, Viral - Blooden_US
dc.subject.meshAntigens, Surface - Genetics - Immunology - Metabolismen_US
dc.subject.meshBlotting, Westernen_US
dc.subject.meshElectrophoresis, Polyacrylamide Gelen_US
dc.subject.meshEnzyme-Linked Immunosorbent Assayen_US
dc.subject.meshHumansen_US
dc.subject.meshImmunoglobulin G - Blooden_US
dc.subject.meshImmunoglobulin M - Blooden_US
dc.subject.meshPlasmids - Geneticsen_US
dc.subject.meshPolymerase Chain Reactionen_US
dc.subject.meshRecombinant Proteins - Immunology - Isolation & Purification - Metabolismen_US
dc.subject.meshSars Virus - Genetics - Immunology - Metabolismen_US
dc.subject.meshSevere Acute Respiratory Syndrome - Blood - Virologyen_US
dc.subject.meshViral Envelope Proteins - Genetics - Immunology - Metabolismen_US
dc.titleCloning, purification, and antigenic characterization of three recombinant fragments derived from SARS-CoV S1 domainen_US
dc.typeArticleen_US
dc.identifier.emailYuen, KY:kyyuen@hkucc.hku.hken_US
dc.identifier.authorityYuen, KY=rp00366en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.pmid16261215-
dc.identifier.scopuseid_2-s2.0-58149229726en_US
dc.identifier.volume19en_US
dc.identifier.issue3en_US
dc.identifier.spage275en_US
dc.identifier.epage278en_US
dc.identifier.scopusauthoridMei, YB=8231190400en_US
dc.identifier.scopusauthoridLiao, ZY=7203032864en_US
dc.identifier.scopusauthoridWang, YD=9638471800en_US
dc.identifier.scopusauthoridZhang, LY=37086714000en_US
dc.identifier.scopusauthoridXu, H=8401769300en_US
dc.identifier.scopusauthoridYuen, KY=36078079100en_US
dc.identifier.scopusauthoridChe, XY=7005743182en_US

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