The development of blackheart resistant pineapples through genetic engineering

Abstract

We are attempting to control blackheart by inhibiting expression of PPO in genetically engineered pineapple plants. PPO genes have been isolated from various pineapple tissues using PCR with redundant oligonucleotides and a cDNA which is specifically induced in blackheart affected fruit has been cloned. Suitable promoters to drive gene expression in pineapple have been defined and gene constructs suitable for transformation of the crop have been prepared. To generate transgenic pineapple we have established an efficient plant regeneration system based on the production of highly regenerable callus from leaf bases of tissue cultured plantlets. A transformation system based on micro-projectile bombardment of this regenerable callus has been developed. In addition, we have also transformed pineapple using Agrobacterium-mediated gene delivery to leaf bases, from which we have generated stably transformed plantlets. For biolistics we have obtained over a dozen independent transformation events in at least four separate experiments, transformation in some instances has been confirmed by Southerns and shoots have been regenerated from this material. For Agrobacterium, transformed plants have been regenerated and multiple events have been observed from at least four separate experiments.