Abstract:
For the characterization of specific promoters in banana (Musa spp.) a tagging approach was applied. Embryogenic cell suspensions were cocultivated with Agrobacterium containing a promoterless luciferase reporter gene next to a T-DNA border. Transgenic cell colonies were screened for luciferase (LUC) activation between two to four months after transformation and selected candidates were further monitored during the regeneration process. The frequency of baseline luciferase activity ranged between 0.9% and 2.5% with an improved tagging vector containing a codon-optimized luciferase (luc+) gene close to the right T-DNA border. This vector was used for tagging cold-responsive promoters by applying different temperatures between 18°C to 8°C. Out of approximately 16,000 transgenic cell colonies, 106 (0.67%) showed cold-responsive LUC activity with either enhanced or repressed expression. In contrast, many of the 26 regenerated in vitro plantlets (0.16%) exerted specific and localized LUC expression patterns in different tissues. The results will contribute to the isolation of native promoters for genetic modification in this crop.
