Production and Characterization of Transgenic Citrus Plants Carrying p35 Anti-apoptotic Gene

The baculovirus gene p35 is known to be negative regulator of apoptosis. Transgenic plants of Carrizo citrange rootstock cultivar [Citrus sinensis (L.) Osbeck x Poncirus trifoliata (L.) Raf.] expressing the p35 gene were produced. Two transgenic lines (P35-2 and P35-4) contained one copy of the p35 gene in their genomes, and line P35-1 contained two copies of the p35 gene. In all experiments performed in this study, we used clonal plants that were propagated from original transgenic plants and the wild-type through rooting of stem explants. When grown in the absence of stress-inducing factors, transgenic plants did not appear to be phenotypically different from the wild-type plants except for the P35-4 line which was observed to contain 10% less stomata on the abaxial side of the leaves. Leaf senescence on detached branches was significantly slower in all three transgenic lines compared to the wild-type. Etephone treatment demonstrated that transgenic lines were still sensitive to ethylene although leaf abscission from detached branches of P35-2 and P35-4 lines occurred at a slower rate while abscission of leaves from wild-type and P35-1 line was not significantly different. Contrary to our observations on leaf senescence, treatment of leaf explants with increasing concentrations of the herbicide Paraquat, resulted in a similar decay of chlorophyll A in all tested lines. Girdling of plant stems, besides other effects, also caused some root loss in all tested lines. However, the loss was less pronounced in P35-2 and P35-4 lines. Interestingly, the roots of P35-2 line maintained high levels of starch 78 days after being girdled. Response of citrus plants to stem girdling is similar to symptoms of infection with the greening-causing bacteria (Las). Slower rate of root decay following the stem girdling suggests that some of transgenic lines may be able to attenuate the death of root following the infection with Las and we are currently producing more clonal plants to test this hypothesis.