Transgenic tobacco plants carrying the non-structural P3 gene of potato virus A

Transgenic tobacco (Nicotiana tobacum) plants carrying the gene coding for potato virus A (PVA) non-structural P3 protein were prepared by inoculation with Agrobacterium tumefaciens. Seeds from self-pollinated flowers (T-1 generation) were collected. To estimate the effectiveness of vertical transfer of the introduced gene and usefulness of respective plant lines for further experiments, the T-1 generation was characterized by testing its ability to grow in the presence of kanamycin (Km) and by PCR of both neomycin phosphotransferase (nptII) and PVA P3 genes. Eight and ten of 29 lines showed Mendelian segregation of Km-resistant phenotype 3:1 and >= 15:1, respectively, the T-1 of eleven lines showed low Km resistance. Selected PCR-positive lines were tested for the presence of P3 mRNA. In most cases the transgene transcription was dependent on the presence or absence of Km in the plant growth medium. Prepared transgenic plants were furthermore tested for sensitivity to PVA and potato virus Y (PVY) infection. All of them showed identical symptom development as the non-transgenic control plants.