Detection of viral infections by an oligonucleotide microarray.
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.
Sip
Miroslav
M
Department of Laboratory Methods and Medical Technology, Faculty of Health and Social Studies, University of South Bohemia, J Boreckeho 27, 370 11 Ceske Budejovice, Czech Republic. sip@zsf.jcu.cz
Bystricka
Dagmar
D
Kmoch
Stanislav
S
Noskova
Lenka
L
Hartmannova
Hana
H
Dedic
Petr
P
D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010935:Plant Diseases / Q000821:virology*; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D011198:Solanum tuberosum / Q000821:virology*; D014773:Virology / Q000379:methods*
D017931:DNA Primers; D018076:DNA, Complementary; D012367:RNA, Viral