Particle-bombardment-mediated co-transformation of elite Chinese rice cultivars with genes conferring resistance to bacterial blight and sap-sucking insect pests

Transgenic rice plants were generated using particle bombardment to simulta neously introduce the Xa21 gene effective against bacterial blight and the Galanthus nivalis agglutinin (snowdrop lectin; gna) gene effective against sap-sucking insect pests, specifically the brown plant hopper. Using three plasmids, we cotransformed 5- to 10-d-old, mature seed-derived rice (Oryza sativa L.) callus of two elite Chinese rice cultivars, Eyi 105 and Ewan 5. The plasmids carried a total of four genes. The gna and Xa21 genes were car ried on separate plasmids. The selectable marker hygromycin phosphotransfer ase (hpt) and the reporter gene beta-glucuronidase (gusA) were linked on th e same, co-integrate vector. We recovered over 160 independently derived tr ansgenic rice plants. Over 70% of the transgenic plants carried all four ge nes, as confirmed by polymerase chain reaction and/or Southern blot analysi s. Furthermore, 70% of transgenic plants carrying all four genes also coexp ressed all four genes, as confirmed by growth on selection media (hpt), GUS histochemical assays (gusA), western blotting (gna) and reverse transcript ase-polymerase chain reaction (Xa21) analysis. The co-expression efficiency reported for the four transgenes in our study is the highest ever found in any transgenic plant population generated through co-transformation. The l inked genes (hpt and gusA) co-integrated with a frequency of near 100%, and we observed a co-integration frequency greater than 70% for the genes carr ied on separate plasmids. We observed no preferential integration of any pa rticular gene(s), Genetic analysis confirmed Mendelian segregation of the t ransgenes in subsequent generations. We report, for the first time, generat ion and analysis of transgenic rice lines carrying genes effective against more than one taxa of pathogen or pest, substantiating that particle bombar dment represents an effective way to introduce unlinked complex multiple tr aits into plants.