Improvement of Agrobacterium-mediated transformation efficiency and transgenic plant regeneration of Vitis vinifera L. by optimizing selection regimes and utilizing cryopreserved cell suspensions

Various concentrations of kanamycin and paromomycin were examined for their effect on embryogenic cell suspension viability, transformation efficiency, and transgenic plant regeneration. Paromomycin (10-25 mg/1) induced an earlier killing effect on cell suspensions than kanamycin (40-100 mg/l). Transformation efficiency and the number of embryos developed on selection medium were positively correlated with an increase in paromomycin concentrations from 10 to 20 or 25 mg/l. Paromomycin was more effective than kanamycin in selection of transformed cells and induction of embryo development during selection. Although no differences in transformation efficiency were observed between cryopreserved and non-cryopreserved cells, conversions of transformed cells to embryos and plantlets were significantly improved when cryopreserved cells were used as the target material. With the improved transformation procedure proposed, 65-80% of GUS-positive cells and embryos were produced with about 40% of transformed embryos regenerated into plants. (C) 2004 Elsevier Ireland Ltd. All rights reserved.