Genetic engineering of cabbage and watercress with B.T.cry genes for insect resistance

The main goal of the research was to develop transgenic cabbage ( Brassica oleracea var. capitata) and watercress (Rorippa nasturtium-aquaticum) transformed with Bacillus thuringiensis (Bt) insecticidal genes for the control of the diamondback moth (Plutella xylostella). Protocols for tissue culture and Agrobacterium-mediated transformation of both cabbage and watercress were developed after many factors that could affect the virulence of Agrobacterium and the regeneration of transformed cells were examined. Factors important for transformation included preculture and coculture of explants on a callus induction medium, induction of Agrobacterium virulence with a minimal medium containing acetosyringone, use of an appropriate amount and a delay in initial application of selective agents. All cabbage plants transgenic for cry1Ab3 provided 100% mortality of the larvae of the diamondback moth, whereas cabbage and watercress plants transformed with cry1Ia3 were not resistant to the moth. Northern analysis showed that while cry1Ab3 plants produced a full-length transcript of the gene, cry1Ia3 plants produced a shortened transcript. We have also obtained transgenic watercress plants transformed with cry1Ab3 and are currently performing molecular characterization and insect bioassay of these plants.; In order to obtain temporal control of the expression of Bt insecticidal genes in transgenic plants, three inducible promoters (PR- 1a from tobacco, vspB from soybean, and pin2 from potato) were cloned and fused with a Bt gene ( cry1Ab3) for plant transformation. Both transgenic tobacco and cabbage plants were obtained that were transformed with the Bt gene under the transcriptional control of any one of the three promoters. Inducible gene expression analysis based on Northern hybridization and insect bioassay showed that while there was constitutive Bt expression in vspB/cry1Ab3 or pin2/cry1Ab3 transgenic plants in the absence of any induction, Bt expression could be tightly controlled with PR-1a/cry1Ab3 plants by salicylic acid induction: there was no expression without induction, high expression occurred in response to induction, and expression declined when induction was removed. Such controllable expression of Bt genes in transgenic plants may turn out to be very useful in delaying development of insect resistance to Bt-transgenic plants.