Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis
Autor(a) principal: | |
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Data de Publicação: | 2004 |
Outros Autores: | , , , |
Tipo de documento: | Relatório |
Idioma: | eng |
Título da fonte: | Fitopatologia Brasileira |
Texto Completo: | http://old.scielo.br/scielo.php?script=sci_arttext&pid=S0100-41582004000500017 |
Resumo: | The coat protein gene of Apple stem grooving virus (ASGV) was amplified by RT-PCR, cloned, sequenced and subcloned in the expression vector pMal-c2. This plasmid was used to transform Escherichia coli BL21c+ competent cells. The ASGV coat protein (cp) was expressed as a fusion protein containing a fragment of E. coli maltose binding protein (MBP). Bacterial cells were disrupted by sonication and the ASGVcp/MBP fusion protein was purified by amylose resin affinity chromatography. Polyclonal antibodies from rabbits immunized with the fusion protein gave specific reactions to ASGV from infected apple (Malus domestica) cv. Fuji Irradiada and Chenopodium quinoa at dilutions of up to 1:1,000 and 1:2,000, respectively, in plate trapped ELISA. The ASGVcp/MBP fusion protein reacted to a commercial antiserum against ASGV in immunoblotting assay. The IgG against ASGVcp/MBP performed favorably in specificity and sensitivity to the virus. This method represents an additional tool for the efficient ASGV-indexing of apple propagative and mother stock materials, and for use in support of biological and molecular techniques. |
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Fitopatologia Brasileira |
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Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosisrecombinant antigenlatent virusELISAMalus domesticaThe coat protein gene of Apple stem grooving virus (ASGV) was amplified by RT-PCR, cloned, sequenced and subcloned in the expression vector pMal-c2. This plasmid was used to transform Escherichia coli BL21c+ competent cells. The ASGV coat protein (cp) was expressed as a fusion protein containing a fragment of E. coli maltose binding protein (MBP). Bacterial cells were disrupted by sonication and the ASGVcp/MBP fusion protein was purified by amylose resin affinity chromatography. Polyclonal antibodies from rabbits immunized with the fusion protein gave specific reactions to ASGV from infected apple (Malus domestica) cv. Fuji Irradiada and Chenopodium quinoa at dilutions of up to 1:1,000 and 1:2,000, respectively, in plate trapped ELISA. The ASGVcp/MBP fusion protein reacted to a commercial antiserum against ASGV in immunoblotting assay. The IgG against ASGVcp/MBP performed favorably in specificity and sensitivity to the virus. This method represents an additional tool for the efficient ASGV-indexing of apple propagative and mother stock materials, and for use in support of biological and molecular techniques.Sociedade Brasileira de Fitopatologia2004-10-01info:eu-repo/semantics/reportinfo:eu-repo/semantics/publishedVersiontext/htmlhttp://old.scielo.br/scielo.php?script=sci_arttext&pid=S0100-41582004000500017Fitopatologia Brasileira v.29 n.5 2004reponame:Fitopatologia Brasileirainstname:Sociedade Brasileira de Fitopatologia (SBF)instacron:SBF10.1590/S0100-41582004000500017info:eu-repo/semantics/openAccessNickel,OsmarTargon,Maria L.P.N.Fajardo,Thor V.M.Machado,Marcos A.Trivilin,Ana P.eng2004-10-25T00:00:00Zoai:scielo:S0100-41582004000500017Revistahttp://www.scielo.br/fbONGhttps://old.scielo.br/oai/scielo-oai.php||sbf-revista@ufla.br1678-46770100-4158opendoar:2004-10-25T00:00Fitopatologia Brasileira - Sociedade Brasileira de Fitopatologia (SBF)false |
dc.title.none.fl_str_mv |
Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis |
title |
Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis |
spellingShingle |
Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis Nickel,Osmar recombinant antigen latent virus ELISA Malus domestica |
title_short |
Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis |
title_full |
Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis |
title_fullStr |
Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis |
title_full_unstemmed |
Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis |
title_sort |
Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis |
author |
Nickel,Osmar |
author_facet |
Nickel,Osmar Targon,Maria L.P.N. Fajardo,Thor V.M. Machado,Marcos A. Trivilin,Ana P. |
author_role |
author |
author2 |
Targon,Maria L.P.N. Fajardo,Thor V.M. Machado,Marcos A. Trivilin,Ana P. |
author2_role |
author author author author |
dc.contributor.author.fl_str_mv |
Nickel,Osmar Targon,Maria L.P.N. Fajardo,Thor V.M. Machado,Marcos A. Trivilin,Ana P. |
dc.subject.por.fl_str_mv |
recombinant antigen latent virus ELISA Malus domestica |
topic |
recombinant antigen latent virus ELISA Malus domestica |
description |
The coat protein gene of Apple stem grooving virus (ASGV) was amplified by RT-PCR, cloned, sequenced and subcloned in the expression vector pMal-c2. This plasmid was used to transform Escherichia coli BL21c+ competent cells. The ASGV coat protein (cp) was expressed as a fusion protein containing a fragment of E. coli maltose binding protein (MBP). Bacterial cells were disrupted by sonication and the ASGVcp/MBP fusion protein was purified by amylose resin affinity chromatography. Polyclonal antibodies from rabbits immunized with the fusion protein gave specific reactions to ASGV from infected apple (Malus domestica) cv. Fuji Irradiada and Chenopodium quinoa at dilutions of up to 1:1,000 and 1:2,000, respectively, in plate trapped ELISA. The ASGVcp/MBP fusion protein reacted to a commercial antiserum against ASGV in immunoblotting assay. The IgG against ASGVcp/MBP performed favorably in specificity and sensitivity to the virus. This method represents an additional tool for the efficient ASGV-indexing of apple propagative and mother stock materials, and for use in support of biological and molecular techniques. |
publishDate |
2004 |
dc.date.none.fl_str_mv |
2004-10-01 |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/report |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
format |
report |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S0100-41582004000500017 |
url |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S0100-41582004000500017 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
10.1590/S0100-41582004000500017 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
text/html |
dc.publisher.none.fl_str_mv |
Sociedade Brasileira de Fitopatologia |
publisher.none.fl_str_mv |
Sociedade Brasileira de Fitopatologia |
dc.source.none.fl_str_mv |
Fitopatologia Brasileira v.29 n.5 2004 reponame:Fitopatologia Brasileira instname:Sociedade Brasileira de Fitopatologia (SBF) instacron:SBF |
instname_str |
Sociedade Brasileira de Fitopatologia (SBF) |
instacron_str |
SBF |
institution |
SBF |
reponame_str |
Fitopatologia Brasileira |
collection |
Fitopatologia Brasileira |
repository.name.fl_str_mv |
Fitopatologia Brasileira - Sociedade Brasileira de Fitopatologia (SBF) |
repository.mail.fl_str_mv |
||sbf-revista@ufla.br |
_version_ |
1754734649885589504 |