Abstract: The bifunctional OvBAN/bar gene was inserted into cell lines of the alfalfa cultivar ‘Gannong No.3’ using Agrobacterium-mediated transformation. PCR and southern blot analysis showed that four transgenic lines that contained the target gene were obtained, and all showed hybridization bands; two lines had single copies, one had two copies, and one had three copies. Expression of OvBAN in the four transgenic plants was analyzed by quantitative RT-PCR, and confirmed to be most highly expressed in the line with 3 copies, and also highly expressed in the two lines with two copies. In these transgenic alfalfa plants, anthocyanin reductase activity and condensed tannin content were significantly higher than in untransformed alfalfa. These three lines of transgenic alfalfa over-expressing the OvBAN/bar gene were tested for resistance to aphids, and the transgenic material demonstrated stronger aphid resistance than non-transgenic plants, with a 78%-93% reduction in aphid numbers per plant. Transgenic plants were then sprayed 0.5% glufosinate to test for herbicide resistance. After 10 days, the transgenic plants were still healthy, while the untransformed plants withered and eventually died, indicating that the bar gene had been integrated into the genome and stably expressed. In summary, the study indicated that introduction of the OvBAN/bar bifunctional genes significantly enhanced insect resistance and herbicide tolerance in transgenic alfalfa plants.

Key words: transgenic alfalfa, OvBAN, bar, insect resistance, herbicide tolerance