TOBACCO (NICOTIANA BENTHAMIANA DOMIN) PLANT TRANSFORMATION WITH AND WITHOUT SELECTABLE MARKER GENE FOR MULTIPLE VIRUS RESISTANCE

Abstract

Transformation of tobacco plants (Nicotiana benthamiana Domin), containing non-translatable coat protein (CP) gene fragments of Potato virus Y (PVY), Potato leafroll virus (PLRV) and Potato virus X (PVX) with and without selectable marker gene was conducted using Agrobacterium-mediated transformation. Preliminary identifications of the transformants were done based on the expression of the reporter gene, Beta-Glucuronidase (GUS) gene. GUS activity test was conducted in the first (T0) generation while the plants were in test tubes and after transferring to a greenhouse for marker-free and marker-aided transformations, respectively. In the transformation with marker gene, three lines, which later on (in T1 generation) became virus resistant and PCR-positive for the insert, were generated from GUS-negative mother plants. Although regeneration of plants from marker-free transformation was relatively easy, it was not possible to get transgenic plants using histochemical GUS staining selection system