1887

Abstract

Potato virus Y (PVY) full-length cDNA has been found to be refractory to cloning in cells. A full-length 9.7 kb PVY cDNA was obtained by reverse transcription polymerase chain reaction (RT-PCR) from the RNA of PVY (tuber necrotic strain, PVY). Double-stranded DNA fragments were used as primers (ds megaprimers), to include signals for transcription (a cauliflower mosaic virus 35S RNA promoter and a nopaline synthase terminator) in the final PCR product. Biolistic bombardment with a helium particle gun was used to inoculate the amplified product to detached tobacco leaves. Inoculation of tobacco plants with ground inoculated leaves followed by northern blot, ELISA and immuno-electron microscopy demonstrated that the DNA was highly infectious with up to 90% of bombarded leaves containing the virus. This methodology will allow the use of reverse genetics in the study of PVY-plant interactions and will also be useful for obtaining infectious cDNA from other viruses with large RNA genomes.

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/content/journal/jgv/10.1099/0022-1317-77-3-519
1996-03-01
2019-10-19
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http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-77-3-519
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