Development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens
cDNA clones of potato virus X (PVXcp strain), potato virus Y (PVYo strain), potato leaf roll virus (PLRV) and potato spindle tuber viroid (PSTV) were used separately or combined for the detection of the corresponding RNAs in extracts of infected plants. A general method for the rapid preparation of...
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1991
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| Acceso en línea: | https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_01660934_v31_n1_p11_Hopp http://hdl.handle.net/20.500.12110/paper_01660934_v31_n1_p11_Hopp |
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paper:paper_01660934_v31_n1_p11_Hopp2025-07-30T17:54:01Z Development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens Hopp, Horacio Esteban Tozzini, Alejandro Carlos Orman, Betina Ceriani, María Fernanda Saladrigas de Isenring, María Verónica Celnik, Rosana Mara Del Vas, Mariana Mentaberry, Alejandro Néstor complementary dna article dna probe dna rna hybridization nonhuman plant virus potato priority journal rna virus virus detection Cloning, Molecular DNA Probes Enzyme-Linked Immunosorbent Assay Methods Nucleic Acid Hybridization Plant Diseases Plant Viruses Potatoes RNA, Viral Sensitivity and Specificity Support, Non-U.S. Gov't Hexapoda Insecta Perodicticus potto Potato leafroll virus Potato spindle tuber viroid Potato virus Y RNA viruses Solanum tuberosum Viroids cDNA clones of potato virus X (PVXcp strain), potato virus Y (PVYo strain), potato leaf roll virus (PLRV) and potato spindle tuber viroid (PSTV) were used separately or combined for the detection of the corresponding RNAs in extracts of infected plants. A general method for the rapid preparation of RNA extracts without use of organic solvents (i.e. phenol) was developed for this purpose. Plant extracts from a range of field, artificially inoculated germplasm genotypes, micro-propagated and protoplast samples, as well as vector insect extracts, were dot-blotted onto nylon or nitrocellulose membranes, subjected to sandwich nucleic acid hybridization with non-labelled specific single-stranded DNA probes followed by a biotin-labelled second step hybridization probe. Each probe was virus-specific but not strain-specific. Healthy or non-related plant extracts developed very faint or no signals. Sensitivity was tested by slot-blot hybridization. Detection levels were between 1.5 to 6 pg of viral nucleic acids and between 20 to 50 times more sensitive than standard double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). The assay developed was tested with material that was prepared for processing in the field (combination of fresh sap with extraction solution) and tested under simple laboratory conditions for detection. It was also successfully employed for screening of germplasm for virus resistance, detection of pathogens in vector insects, plantlets grown in vitro and in more sophisticated quantitative determinations of viral replication in artificially inoculated plants and protoplasts. © 1991. Fil:Hopp, H.E. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Tozzini, A.C. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Orman, B. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Ceriani, M.F. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Saladrigas, M.V. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Celnik, R. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:del Vas, M. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Mentaberry, A.N. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. 1991 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_01660934_v31_n1_p11_Hopp http://hdl.handle.net/20.500.12110/paper_01660934_v31_n1_p11_Hopp |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
complementary dna article dna probe dna rna hybridization nonhuman plant virus potato priority journal rna virus virus detection Cloning, Molecular DNA Probes Enzyme-Linked Immunosorbent Assay Methods Nucleic Acid Hybridization Plant Diseases Plant Viruses Potatoes RNA, Viral Sensitivity and Specificity Support, Non-U.S. Gov't Hexapoda Insecta Perodicticus potto Potato leafroll virus Potato spindle tuber viroid Potato virus Y RNA viruses Solanum tuberosum Viroids |
| spellingShingle |
complementary dna article dna probe dna rna hybridization nonhuman plant virus potato priority journal rna virus virus detection Cloning, Molecular DNA Probes Enzyme-Linked Immunosorbent Assay Methods Nucleic Acid Hybridization Plant Diseases Plant Viruses Potatoes RNA, Viral Sensitivity and Specificity Support, Non-U.S. Gov't Hexapoda Insecta Perodicticus potto Potato leafroll virus Potato spindle tuber viroid Potato virus Y RNA viruses Solanum tuberosum Viroids Hopp, Horacio Esteban Tozzini, Alejandro Carlos Orman, Betina Ceriani, María Fernanda Saladrigas de Isenring, María Verónica Celnik, Rosana Mara Del Vas, Mariana Mentaberry, Alejandro Néstor Development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens |
| topic_facet |
complementary dna article dna probe dna rna hybridization nonhuman plant virus potato priority journal rna virus virus detection Cloning, Molecular DNA Probes Enzyme-Linked Immunosorbent Assay Methods Nucleic Acid Hybridization Plant Diseases Plant Viruses Potatoes RNA, Viral Sensitivity and Specificity Support, Non-U.S. Gov't Hexapoda Insecta Perodicticus potto Potato leafroll virus Potato spindle tuber viroid Potato virus Y RNA viruses Solanum tuberosum Viroids |
| description |
cDNA clones of potato virus X (PVXcp strain), potato virus Y (PVYo strain), potato leaf roll virus (PLRV) and potato spindle tuber viroid (PSTV) were used separately or combined for the detection of the corresponding RNAs in extracts of infected plants. A general method for the rapid preparation of RNA extracts without use of organic solvents (i.e. phenol) was developed for this purpose. Plant extracts from a range of field, artificially inoculated germplasm genotypes, micro-propagated and protoplast samples, as well as vector insect extracts, were dot-blotted onto nylon or nitrocellulose membranes, subjected to sandwich nucleic acid hybridization with non-labelled specific single-stranded DNA probes followed by a biotin-labelled second step hybridization probe. Each probe was virus-specific but not strain-specific. Healthy or non-related plant extracts developed very faint or no signals. Sensitivity was tested by slot-blot hybridization. Detection levels were between 1.5 to 6 pg of viral nucleic acids and between 20 to 50 times more sensitive than standard double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). The assay developed was tested with material that was prepared for processing in the field (combination of fresh sap with extraction solution) and tested under simple laboratory conditions for detection. It was also successfully employed for screening of germplasm for virus resistance, detection of pathogens in vector insects, plantlets grown in vitro and in more sophisticated quantitative determinations of viral replication in artificially inoculated plants and protoplasts. © 1991. |
| author |
Hopp, Horacio Esteban Tozzini, Alejandro Carlos Orman, Betina Ceriani, María Fernanda Saladrigas de Isenring, María Verónica Celnik, Rosana Mara Del Vas, Mariana Mentaberry, Alejandro Néstor |
| author_facet |
Hopp, Horacio Esteban Tozzini, Alejandro Carlos Orman, Betina Ceriani, María Fernanda Saladrigas de Isenring, María Verónica Celnik, Rosana Mara Del Vas, Mariana Mentaberry, Alejandro Néstor |
| author_sort |
Hopp, Horacio Esteban |
| title |
Development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens |
| title_short |
Development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens |
| title_full |
Development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens |
| title_fullStr |
Development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens |
| title_full_unstemmed |
Development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens |
| title_sort |
development and application of a nonradioactive nucleic acid hybridization system for simultaneous detection of four potato pathogens |
| publishDate |
1991 |
| url |
https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_01660934_v31_n1_p11_Hopp http://hdl.handle.net/20.500.12110/paper_01660934_v31_n1_p11_Hopp |
| work_keys_str_mv |
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| _version_ |
1840322280183300096 |