Genetic Transformation of Embryogenic Cultures and Recovery of Transgenic Plants in Vitis Vinifera, Vitis Rotundifolia and Vitis Hybrids

Abstract

Factors influencing Agrobacterium-mediated transformation of 15 Vitis rotundifolia, Vitis rupestris, Vitis vinifera and Vitis hybrids were explored. A green fluorescent protein/neomycin phosphotransferase II (GFP/NPT II) fusion gene was used to measure transient and stable transgene expression levels. Embryogenic cell cultures at different developmental stages were co-cultivated with Agrobacterium to determine the best stage for transformation. The effect of antioxidant treatment on recovery of transgenic embryo and plant lines was also studied. Transient and stable GFP expression varied widely among the species. For example, 1-81% of V. vinifera somatic embryos exhibited transient GFP expression, depending on the cultivar. Similarly, 0-60% of co-cultivated embryos produced stable transgenic lines. Proembryonal masses and cotyledonary stage somatic embryos were best for transformation of V. rotundifolia, V. rupestris and certain Vitis hybrids, whereas only cotyledonary stage somatic embryos were best for transformation of V. vinifera. Agrobacterium-induced browning/ necrosis was reduced by adding 1 g L-1 dithiothreitol (DTT) antioxidant to the post cocultivation wash medium. Transgenic plants have been recovered from V. rupestris 'St.

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