Improved Detection of Citrus psorosis virus and Coat Protein‐Derived Transgenes in Citrus Plants: Comparison Between RT‐qPCR and TAS‐ELISA
Citrus is one of the most economically important fruit crops in the world. Citrus psorosis is a serious disease affecting mainly oranges and mandarins in Argentina and Uruguay. The causal agent is Citrus psorosis virus (CPsV), an ophiovirus with a tripartite ssRNA genome of negative polarity. The co...
| Autores principales: | , , , |
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| Formato: | info:ar-repo/semantics/artículo |
| Lenguaje: | Inglés |
| Publicado: |
Wiley
2019
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| Materias: | |
| Acceso en línea: | https://onlinelibrary.wiley.com/doi/10.1111/jph.12392 http://hdl.handle.net/20.500.12123/4684 https://doi.org/10.1111/jph.12392 |
| _version_ | 1855035365766725632 |
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| author | de Francesco, Agustina Costa, Norma Beatriz Plata Tamayo, Maria Ines Garcia, Maria Laura |
| author_browse | Costa, Norma Beatriz Garcia, Maria Laura Plata Tamayo, Maria Ines de Francesco, Agustina |
| author_facet | de Francesco, Agustina Costa, Norma Beatriz Plata Tamayo, Maria Ines Garcia, Maria Laura |
| author_sort | de Francesco, Agustina |
| collection | INTA Digital |
| description | Citrus is one of the most economically important fruit crops in the world. Citrus psorosis is a serious disease affecting mainly oranges and mandarins in Argentina and Uruguay. The causal agent is Citrus psorosis virus (CPsV), an ophiovirus with a tripartite ssRNA genome of negative polarity. The coat protein (CP), the most abundant viral protein in infected plants, has been used to detect CPsV by TAS‐ELISA, but only biological indexing, requiring 1 year, is the current and validated technique for diagnosis of citrus psorosis. In this study, a SYBR Green RT‐qPCR protocol was developed, with primers designed to the most conserved region of the cp gene. We tested their specificity and sensitivity in comparison with TAS‐ELISA. This RT‐qPCR was applied successfully to field samples from Argentina, to a variety of isolates from different countries maintained in the greenhouse, to young seedlings and old trees from a psorosis natural transmission plot, and to transgenic citrus expressing the cp gene of CPsV or a fragment thereof. This method allowed accurate quantification of viral titer and cp gene expression in transgenic plants, which could not be detected previously. The sensitivity and reliability of quantitative CPsV detection were improved with greater speed using commercial reagents, and the sensitivity was three orders of magnitude higher than that of TAS‐ELISA. All these data encourage its validation. |
| format | info:ar-repo/semantics/artículo |
| id | INTA4684 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2019 |
| publishDateRange | 2019 |
| publishDateSort | 2019 |
| publisher | Wiley |
| publisherStr | Wiley |
| record_format | dspace |
| spelling | INTA46842019-03-20T14:51:40Z Improved Detection of Citrus psorosis virus and Coat Protein‐Derived Transgenes in Citrus Plants: Comparison Between RT‐qPCR and TAS‐ELISA de Francesco, Agustina Costa, Norma Beatriz Plata Tamayo, Maria Ines Garcia, Maria Laura Citrus Enfermedades de las Plantas Virus de las Plantas Proteínas de la Cubierta PCR ELISA Diagnóstico Plant Diseases Plant Viruses Coat Proteins Diagnosis Virus de la Psoriasis de los Cítricos Citrus Psorosis Virus Citrus is one of the most economically important fruit crops in the world. Citrus psorosis is a serious disease affecting mainly oranges and mandarins in Argentina and Uruguay. The causal agent is Citrus psorosis virus (CPsV), an ophiovirus with a tripartite ssRNA genome of negative polarity. The coat protein (CP), the most abundant viral protein in infected plants, has been used to detect CPsV by TAS‐ELISA, but only biological indexing, requiring 1 year, is the current and validated technique for diagnosis of citrus psorosis. In this study, a SYBR Green RT‐qPCR protocol was developed, with primers designed to the most conserved region of the cp gene. We tested their specificity and sensitivity in comparison with TAS‐ELISA. This RT‐qPCR was applied successfully to field samples from Argentina, to a variety of isolates from different countries maintained in the greenhouse, to young seedlings and old trees from a psorosis natural transmission plot, and to transgenic citrus expressing the cp gene of CPsV or a fragment thereof. This method allowed accurate quantification of viral titer and cp gene expression in transgenic plants, which could not be detected previously. The sensitivity and reliability of quantitative CPsV detection were improved with greater speed using commercial reagents, and the sensitivity was three orders of magnitude higher than that of TAS‐ELISA. All these data encourage its validation. EEA Concordia Fil: de Francesco, Agustina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina Fil: Costa, Norma Beatriz. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Concordia; Argentina Fil: Plata Tamayo, Maria Ines. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Concordia; Argentina Fil: Garcia, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina 2019-03-20T14:49:50Z 2019-03-20T14:49:50Z 2015-12 info:ar-repo/semantics/artículo info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion https://onlinelibrary.wiley.com/doi/10.1111/jph.12392 http://hdl.handle.net/20.500.12123/4684 0931-1785 1439-0434 https://doi.org/10.1111/jph.12392 eng info:eu-repo/semantics/restrictedAccess application/pdf Wiley Journal of Phytopathology 163 (11-12) : 915-925 (December 2015) |
| spellingShingle | Citrus Enfermedades de las Plantas Virus de las Plantas Proteínas de la Cubierta PCR ELISA Diagnóstico Plant Diseases Plant Viruses Coat Proteins Diagnosis Virus de la Psoriasis de los Cítricos Citrus Psorosis Virus de Francesco, Agustina Costa, Norma Beatriz Plata Tamayo, Maria Ines Garcia, Maria Laura Improved Detection of Citrus psorosis virus and Coat Protein‐Derived Transgenes in Citrus Plants: Comparison Between RT‐qPCR and TAS‐ELISA |
| title | Improved Detection of Citrus psorosis virus and Coat Protein‐Derived Transgenes in Citrus Plants: Comparison Between RT‐qPCR and TAS‐ELISA |
| title_full | Improved Detection of Citrus psorosis virus and Coat Protein‐Derived Transgenes in Citrus Plants: Comparison Between RT‐qPCR and TAS‐ELISA |
| title_fullStr | Improved Detection of Citrus psorosis virus and Coat Protein‐Derived Transgenes in Citrus Plants: Comparison Between RT‐qPCR and TAS‐ELISA |
| title_full_unstemmed | Improved Detection of Citrus psorosis virus and Coat Protein‐Derived Transgenes in Citrus Plants: Comparison Between RT‐qPCR and TAS‐ELISA |
| title_short | Improved Detection of Citrus psorosis virus and Coat Protein‐Derived Transgenes in Citrus Plants: Comparison Between RT‐qPCR and TAS‐ELISA |
| title_sort | improved detection of citrus psorosis virus and coat protein derived transgenes in citrus plants comparison between rt qpcr and tas elisa |
| topic | Citrus Enfermedades de las Plantas Virus de las Plantas Proteínas de la Cubierta PCR ELISA Diagnóstico Plant Diseases Plant Viruses Coat Proteins Diagnosis Virus de la Psoriasis de los Cítricos Citrus Psorosis Virus |
| url | https://onlinelibrary.wiley.com/doi/10.1111/jph.12392 http://hdl.handle.net/20.500.12123/4684 https://doi.org/10.1111/jph.12392 |
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