The success of DNA expression microarrays has been followed by applications of this technology to molecular diagnosis, mainly in the fields of biology and medicine. The experiments described below apply microarray diagnosis to agriculture. This report presents results of field tests for a DNA microarray designed to diagnose major viral potato pathogens. The assays were performed on samples that had been tested previously for the presence of viral infection by ELISA. RNA isolation methods were optimised for high sensitivity, using only 3 microg of total RNA that were reverse transcribed using random hexamers, with the resulting cDNA hybridised after labelling to an oligonucleotide array. The results obtained confirm the presence of pathogens indicated by ELISA and simultaneously reveal other viruses in the same reaction, showing that this method is appropriate for rapid detection of mixed viral infections. This observation was verified by subsequent RT-PCR and sequencing.

• MeSH
• DNA primery MeSH
• ELISA metody   MeSH
• hybridizace nukleových kyselin MeSH
• komplementární DNA genetika   izolace a purifikace   MeSH
• nemoci rostlin virologie   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• RNA virová genetika   izolace a purifikace   MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů metody   MeSH
• Solanum tuberosum virologie   MeSH
• virologie metody   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• validační studie MeSH

The gene encoding the coat protein (CP) of a potato virus Y (PVY) was cloned into expression vector pMPM-A4Omega. PVY CP was expressed in Escherichia coli and the purified recombinant protein was used for raising rabbit polyclonal antibodies. The sera and antibodies were tested for the detection of PVY in the laboratory host Nicotiana tabacum cv. Petit Havana SR1 and in various cultivars of the natural host Solanum tuberosum by ELISA as well as by Western blots. The antibodies can be used for the detection of the whole strain spectrum of PVY by indirect plate trapped antigen ELISA and Western blot, but not by double antigen sandwich ELISA.

• MeSH
• ELISA MeSH
• králíci MeSH
• monoklonální protilátky biosyntéza   MeSH
• nemoci rostlin virologie   MeSH
• Potyvirus imunologie   MeSH
• protilátky virové biosyntéza   MeSH
• rekombinantní proteiny imunologie   MeSH
• Solanum tuberosum virologie   MeSH
• tabák virologie   MeSH
• virové plášťové proteiny imunologie   MeSH
• western blotting MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• zvířata MeSH