A multiplex qPCR TaqMan-assay to detect fungal antagonism between Trichoderma atroviride (Hypocreaceae) and Botrytis cinerea (Sclerotiniaceae) in blackberry fruits using a de novo tef1-α- and an IGS-sequence based probes
The aim of this study was to design a Trichoderma atroviride-specific qPCR oligo set, evaluate its specificity, and standardize a methodology that quantifies antagonism against Botrytis cinerea in blackberry fruits (Rubus adenotrichos Schltdl.). Primers and probe were designed based on the nuclear t...
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| Format: | Article |
| Language: | English |
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Elsevier
2020-09-01
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| Series: | Biotechnology Reports |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2215017X19305454 |
| _version_ | 1818403183403204608 |
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| author | Irena Hilje-Rodríguez Federico J. Albertazzi German Rivera-Coto Ramón Molina-Bravo |
| author_facet | Irena Hilje-Rodríguez Federico J. Albertazzi German Rivera-Coto Ramón Molina-Bravo |
| author_sort | Irena Hilje-Rodríguez |
| collection | DOAJ |
| description | The aim of this study was to design a Trichoderma atroviride-specific qPCR oligo set, evaluate its specificity, and standardize a methodology that quantifies antagonism against Botrytis cinerea in blackberry fruits (Rubus adenotrichos Schltdl.). Primers and probe were designed based on the nuclear translation elongation factor 1-alpha (tef1-α) of T. atroviride. A commercial IGS-based oligo set was used to quantify B. cinerea. The specificity of the designed oligo set, along with ITS-based oligo sets, was assessed using other Trichoderma species and B. cinerea. Multiplex qPCR assays were performed using DNA from B. cinerea, T. atroviride, and blackberries inoculated with these fungi. Assays with the tef1-α oligo set showed high sensitivity and reproducibility. In inoculated fruits, T. atroviride and B. cinerea were quantified simultaneously, including in symptomless tissues. This work standardized a qPCR methodology that specifically targets a T. atroviride isolate. This newly-designed qPCR oligo set could be useful in future biological control programs. |
| first_indexed | 2024-12-14T08:20:12Z |
| format | Article |
| id | doaj.art-0f9efeb267094e6f971ccc324fc889e0 |
| institution | Directory Open Access Journal |
| issn | 2215-017X |
| language | English |
| last_indexed | 2024-12-14T08:20:12Z |
| publishDate | 2020-09-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Biotechnology Reports |
| spelling | doaj.art-0f9efeb267094e6f971ccc324fc889e02022-12-21T23:09:48ZengElsevierBiotechnology Reports2215-017X2020-09-0127e00447A multiplex qPCR TaqMan-assay to detect fungal antagonism between Trichoderma atroviride (Hypocreaceae) and Botrytis cinerea (Sclerotiniaceae) in blackberry fruits using a de novo tef1-α- and an IGS-sequence based probesIrena Hilje-Rodríguez0Federico J. Albertazzi1German Rivera-Coto2Ramón Molina-Bravo3Escuela de Ciencias Agrarias, Universidad Nacional, Apartado Postal 86-3000, Heredia, Costa Rica; Corresponding author.Centro de Investigación en Biología Celular y Molecular, Universidad de Costa Rica, Apartado Postal 11501-2060, San José, Costa RicaEscuela de Ciencias Agrarias, Universidad Nacional, Apartado Postal 86-3000, Heredia, Costa RicaEscuela de Ciencias Agrarias, Universidad Nacional, Apartado Postal 86-3000, Heredia, Costa RicaThe aim of this study was to design a Trichoderma atroviride-specific qPCR oligo set, evaluate its specificity, and standardize a methodology that quantifies antagonism against Botrytis cinerea in blackberry fruits (Rubus adenotrichos Schltdl.). Primers and probe were designed based on the nuclear translation elongation factor 1-alpha (tef1-α) of T. atroviride. A commercial IGS-based oligo set was used to quantify B. cinerea. The specificity of the designed oligo set, along with ITS-based oligo sets, was assessed using other Trichoderma species and B. cinerea. Multiplex qPCR assays were performed using DNA from B. cinerea, T. atroviride, and blackberries inoculated with these fungi. Assays with the tef1-α oligo set showed high sensitivity and reproducibility. In inoculated fruits, T. atroviride and B. cinerea were quantified simultaneously, including in symptomless tissues. This work standardized a qPCR methodology that specifically targets a T. atroviride isolate. This newly-designed qPCR oligo set could be useful in future biological control programs.http://www.sciencedirect.com/science/article/pii/S2215017X19305454Biological controlMolecular quantificationHydrolysis probeqPCRRubus adenotrichos |
| spellingShingle | Irena Hilje-Rodríguez Federico J. Albertazzi German Rivera-Coto Ramón Molina-Bravo A multiplex qPCR TaqMan-assay to detect fungal antagonism between Trichoderma atroviride (Hypocreaceae) and Botrytis cinerea (Sclerotiniaceae) in blackberry fruits using a de novo tef1-α- and an IGS-sequence based probes Biotechnology Reports Biological control Molecular quantification Hydrolysis probe qPCR Rubus adenotrichos |
| title | A multiplex qPCR TaqMan-assay to detect fungal antagonism between Trichoderma atroviride (Hypocreaceae) and Botrytis cinerea (Sclerotiniaceae) in blackberry fruits using a de novo tef1-α- and an IGS-sequence based probes |
| title_full | A multiplex qPCR TaqMan-assay to detect fungal antagonism between Trichoderma atroviride (Hypocreaceae) and Botrytis cinerea (Sclerotiniaceae) in blackberry fruits using a de novo tef1-α- and an IGS-sequence based probes |
| title_fullStr | A multiplex qPCR TaqMan-assay to detect fungal antagonism between Trichoderma atroviride (Hypocreaceae) and Botrytis cinerea (Sclerotiniaceae) in blackberry fruits using a de novo tef1-α- and an IGS-sequence based probes |
| title_full_unstemmed | A multiplex qPCR TaqMan-assay to detect fungal antagonism between Trichoderma atroviride (Hypocreaceae) and Botrytis cinerea (Sclerotiniaceae) in blackberry fruits using a de novo tef1-α- and an IGS-sequence based probes |
| title_short | A multiplex qPCR TaqMan-assay to detect fungal antagonism between Trichoderma atroviride (Hypocreaceae) and Botrytis cinerea (Sclerotiniaceae) in blackberry fruits using a de novo tef1-α- and an IGS-sequence based probes |
| title_sort | multiplex qpcr taqman assay to detect fungal antagonism between trichoderma atroviride hypocreaceae and botrytis cinerea sclerotiniaceae in blackberry fruits using a de novo tef1 α and an igs sequence based probes |
| topic | Biological control Molecular quantification Hydrolysis probe qPCR Rubus adenotrichos |
| url | http://www.sciencedirect.com/science/article/pii/S2215017X19305454 |
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