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Title: Development of molecular resistance in potato against potato leaf roll virus and potato virus Y through Agrobacterium-mediated double transgenesis

Author
item ARIF, M - Agricultural University Peshawar
item THOMAS, PETER - Retired ARS Employee
item Crosslin, James
item Brown, Charles

Submitted to: Pakistan Journal of Botany
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/1/2008
Publication Date: 4/1/2009
Citation: Arif, M., Thomas, P.E., Crosslin, J., Brown, C.R. 2009. Development of molecular resistance in potato against potato leaf roll virus and potato virus Y through Agrobacterium-mediated double transgenesis. Pakistan Journal of Botany. 41:945-954.

Interpretive Summary: Potato leafroll virus (PLRV) and Potato virus Y (PVY) are the two most economically important viruses of cultivated potato worldwide. This paper reports the production of genetically engineered potatoes of cultivar 'Desiree' with genes designed to confer resistance to both PLRV and PVY. Cultivar Desiree is particularly high-yielding in many regions of the world and virus resistance in this cultivar is of particular interest in some developing nations because of the inability to chemically aphids, which transmit both PLRV and PVY.

Technical Abstract: Potato leafroll virus (PLRV) and potato virus Y (PVY) are the two major viral problems for the potato production all over the world. Transgenic approaches involving the expression of viral genes are being developed to provide protection for plants against viral diseases. The purpose of this study was to develop double transgenic plants of potato using PLRV replicase and PVY coat protein genes tandemly placed in a single T-DNA transformant through Agrobacterium-mediated transformation. A total of 17 lines of putative transformants of potato cv. Desiree were generated from kanamycine resistant calli originated from co-inoculation of separate Agrobacterium cultures containing PVY CP and PLRV replicase genes. Shoots were excised and cultured onto shoot medium containing 250mg/L cefotaxime and 50mg/L kanamycin sulfate in test tubes. Polymerase chain reaction (PCR) analysis was conducted of 39 plants of 16 transformed lines using primers each of PVY CP and PLRV-replicase genes; 10 plants of 8 lines and 7 plants of 6 lines showed presence of of PVY CP and PLRV-replicase genes, respectively. However, 22 plants of 14 lines harbored both PVY-CP (508bp) and PLRV-replicase (449bp) genes. Sixteen plants of 11 double transgenic lines that showed high level of expression of both PLRV-replicase and PVY-CP genes. Transformants and control standards were exposed to field virus infection augmented by placement of aphids on Datura leaves infected with PLRV. Two clones (Des (CP+LR) 9.1 and Des (CP + LR) 9.2) showed incidence of infection statistically lower than the lowest infection of one of the standards (Ranger). The Monsanto clone (21-350) showed no infection. The two resistant clones may be identical as they were derived from the same callus.