Reference gene selection for gene expression studies using RT-qPCR in virus-infected planthoppers
Identifieur interne : 000C31 ( Main/Exploration ); précédent : 000C30; suivant : 000C32Reference gene selection for gene expression studies using RT-qPCR in virus-infected planthoppers
Auteurs : Guillermo A. Maroniche [Argentine] ; M Nica Sagadín [Argentine] ; Vanesa C. Mongelli [Argentine] ; Graciela A. Truol [Argentine] ; Mariana Del Vas [Argentine]Source :
- Virology Journal [ 1743-422X ] ; 2011.
Abstract
Planthoppers not only severely affect crops by causing mechanical damage when feeding but are also vectors of several plant virus species. The analysis of gene expression in persistently infected planthoppers might unveil the molecular basis of viral transmission. Quantitative real-time RT-PCR (RT-qPCR) is currently the most accurate and sensitive method used for quantitative gene expression analysis. In order to normalize the resulting quantitative data, reference genes with constant expression during the experimental procedures are needed.
Partial sequences of the commonly used reference genes actin (ACT), α1-tubulin (TUB), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), elongation factor 1 alpha (EF1A), ribosomal protein S18 (RPS18) and polyubiquitin C (UBI) from
A RT-qPCR platform for gene expression analysis in the MRCV-infected planthopper vector
Url:
DOI: 10.1186/1743-422X-8-308
PubMed: 21679431
PubMed Central: 3142240
Affiliations:
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<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Reference gene selection for gene expression studies using RT-qPCR in virus-infected planthoppers</title><author><name sortKey="Maroniche, Guillermo A" sort="Maroniche, Guillermo A" uniqKey="Maroniche G" first="Guillermo A" last="Maroniche">Guillermo A. Maroniche</name><affiliation wicri:level="1"><nlm:aff id="I1">Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires</wicri:regionArea><wicri:noRegion>Buenos Aires</wicri:noRegion></affiliation></author><author><name sortKey="Sagadin, M Nica" sort="Sagadin, M Nica" uniqKey="Sagadin M" first="M Nica" last="Sagadín">M Nica Sagadín</name><affiliation wicri:level="1"><nlm:aff id="I2">Instituto de Fitopatología y Fisiología Vegetal, Instituto Nacional de Tecnología Agropecuaria (IFFIVE-INTA), Camino 60 cuadras km 5 1/2, X5020ICA, Córdoba, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Fitopatología y Fisiología Vegetal, Instituto Nacional de Tecnología Agropecuaria (IFFIVE-INTA), Camino 60 cuadras km 5 1/2, X5020ICA, Córdoba</wicri:regionArea><wicri:noRegion>Córdoba</wicri:noRegion></affiliation></author><author><name sortKey="Mongelli, Vanesa C" sort="Mongelli, Vanesa C" uniqKey="Mongelli V" first="Vanesa C" last="Mongelli">Vanesa C. Mongelli</name><affiliation wicri:level="1"><nlm:aff id="I1">Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires</wicri:regionArea><wicri:noRegion>Buenos Aires</wicri:noRegion></affiliation></author><author><name sortKey="Truol, Graciela A" sort="Truol, Graciela A" uniqKey="Truol G" first="Graciela A" last="Truol">Graciela A. Truol</name><affiliation wicri:level="1"><nlm:aff id="I2">Instituto de Fitopatología y Fisiología Vegetal, Instituto Nacional de Tecnología Agropecuaria (IFFIVE-INTA), Camino 60 cuadras km 5 1/2, X5020ICA, Córdoba, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Fitopatología y Fisiología Vegetal, Instituto Nacional de Tecnología Agropecuaria (IFFIVE-INTA), Camino 60 cuadras km 5 1/2, X5020ICA, Córdoba</wicri:regionArea><wicri:noRegion>Córdoba</wicri:noRegion></affiliation></author><author><name sortKey="Del Vas, Mariana" sort="Del Vas, Mariana" uniqKey="Del Vas M" first="Mariana" last="Del Vas">Mariana Del Vas</name><affiliation wicri:level="1"><nlm:aff id="I1">Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires</wicri:regionArea><wicri:noRegion>Buenos Aires</wicri:noRegion></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">21679431</idno><idno type="pmc">3142240</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3142240</idno><idno type="RBID">PMC:3142240</idno><idno type="doi">10.1186/1743-422X-8-308</idno><date when="2011">2011</date><idno type="wicri:Area/Pmc/Corpus">000734</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000734</idno><idno type="wicri:Area/Pmc/Curation">000734</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Curation">000734</idno><idno type="wicri:Area/Pmc/Checkpoint">000541</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Checkpoint">000541</idno><idno type="wicri:Area/Ncbi/Merge">000227</idno><idno type="wicri:Area/Ncbi/Curation">000227</idno><idno type="wicri:Area/Ncbi/Checkpoint">000227</idno><idno type="wicri:Area/Main/Merge">000C33</idno><idno type="wicri:Area/Main/Curation">000C31</idno><idno type="wicri:Area/Main/Exploration">000C31</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Reference gene selection for gene expression studies using RT-qPCR in virus-infected planthoppers</title><author><name sortKey="Maroniche, Guillermo A" sort="Maroniche, Guillermo A" uniqKey="Maroniche G" first="Guillermo A" last="Maroniche">Guillermo A. Maroniche</name><affiliation wicri:level="1"><nlm:aff id="I1">Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires</wicri:regionArea><wicri:noRegion>Buenos Aires</wicri:noRegion></affiliation></author><author><name sortKey="Sagadin, M Nica" sort="Sagadin, M Nica" uniqKey="Sagadin M" first="M Nica" last="Sagadín">M Nica Sagadín</name><affiliation wicri:level="1"><nlm:aff id="I2">Instituto de Fitopatología y Fisiología Vegetal, Instituto Nacional de Tecnología Agropecuaria (IFFIVE-INTA), Camino 60 cuadras km 5 1/2, X5020ICA, Córdoba, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Fitopatología y Fisiología Vegetal, Instituto Nacional de Tecnología Agropecuaria (IFFIVE-INTA), Camino 60 cuadras km 5 1/2, X5020ICA, Córdoba</wicri:regionArea><wicri:noRegion>Córdoba</wicri:noRegion></affiliation></author><author><name sortKey="Mongelli, Vanesa C" sort="Mongelli, Vanesa C" uniqKey="Mongelli V" first="Vanesa C" last="Mongelli">Vanesa C. Mongelli</name><affiliation wicri:level="1"><nlm:aff id="I1">Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires</wicri:regionArea><wicri:noRegion>Buenos Aires</wicri:noRegion></affiliation></author><author><name sortKey="Truol, Graciela A" sort="Truol, Graciela A" uniqKey="Truol G" first="Graciela A" last="Truol">Graciela A. Truol</name><affiliation wicri:level="1"><nlm:aff id="I2">Instituto de Fitopatología y Fisiología Vegetal, Instituto Nacional de Tecnología Agropecuaria (IFFIVE-INTA), Camino 60 cuadras km 5 1/2, X5020ICA, Córdoba, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Fitopatología y Fisiología Vegetal, Instituto Nacional de Tecnología Agropecuaria (IFFIVE-INTA), Camino 60 cuadras km 5 1/2, X5020ICA, Córdoba</wicri:regionArea><wicri:noRegion>Córdoba</wicri:noRegion></affiliation></author><author><name sortKey="Del Vas, Mariana" sort="Del Vas, Mariana" uniqKey="Del Vas M" first="Mariana" last="Del Vas">Mariana Del Vas</name><affiliation wicri:level="1"><nlm:aff id="I1">Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires, Argentina</nlm:aff><country xml:lang="fr">Argentine</country><wicri:regionArea>Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (IB-INTA), Las Cabañas y Los Reseros s/n, Hurlingham CP 1686, Buenos Aires</wicri:regionArea><wicri:noRegion>Buenos Aires</wicri:noRegion></affiliation></author></analytic><series><title level="j">Virology Journal</title><idno type="eISSN">1743-422X</idno><imprint><date when="2011">2011</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><sec><title>Background</title><p>Planthoppers not only severely affect crops by causing mechanical damage when feeding but are also vectors of several plant virus species. The analysis of gene expression in persistently infected planthoppers might unveil the molecular basis of viral transmission. Quantitative real-time RT-PCR (RT-qPCR) is currently the most accurate and sensitive method used for quantitative gene expression analysis. In order to normalize the resulting quantitative data, reference genes with constant expression during the experimental procedures are needed.</p></sec><sec><title>Results</title><p>Partial sequences of the commonly used reference genes actin (ACT), α1-tubulin (TUB), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), elongation factor 1 alpha (EF1A), ribosomal protein S18 (RPS18) and polyubiquitin C (UBI) from <italic>Delphacodes kuscheli</italic>, a planthopper capable of persistently transmitting the plant fijivirus <italic>Mal de Río Cuarto virus </italic>(MRCV), were isolated for the first time. Specific RT-qPCR primers were designed and the expression stability of these genes was assayed in MRCV-infective and naïve planthoppers using geNorm, Normfinder and BestKeeper tools. The overall analysis showed that UBI, followed by 18S and ACT, are the most suitable genes as internal controls for quantitative gene expression studies in MRCV-infective planthoppers, while TUB and EF1A are the most variable ones. Moreover, EF1A was upregulated by MRCV infection.</p></sec><sec><title>Conclusions</title><p>A RT-qPCR platform for gene expression analysis in the MRCV-infected planthopper vector <italic>Delphacodes kuscheli </italic>was developed. 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last="Maroniche">Guillermo A. Maroniche</name></noRegion><name sortKey="Del Vas, Mariana" sort="Del Vas, Mariana" uniqKey="Del Vas M" first="Mariana" last="Del Vas">Mariana Del Vas</name><name sortKey="Mongelli, Vanesa C" sort="Mongelli, Vanesa C" uniqKey="Mongelli V" first="Vanesa C" last="Mongelli">Vanesa C. Mongelli</name><name sortKey="Sagadin, M Nica" sort="Sagadin, M Nica" uniqKey="Sagadin M" first="M Nica" last="Sagadín">M Nica Sagadín</name><name sortKey="Truol, Graciela A" sort="Truol, Graciela A" uniqKey="Truol G" first="Graciela A" last="Truol">Graciela A. Truol</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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