Multiple Shoot Induction from Seedling Epicotyls and Transgenic Citrus Plant Regeneration Containing the Green Fluorescent Protein Gene

This research aimed to optimize the organogenesis of epicotyl segments and to efficiently obtain transgenic plants of 'Bingtang' sweet orange (Citrus sinensis L. Osb.), an elite citrus cultivar in China. Organogenesis induction was induced in epicotyl segments of 2 weeks old seedlings of this cultivar. Two important factors influencing organogenesis in vitro viz, hormone combination (IAA and BA) and cut modes were characterized. IAA had a positive effect on bud formation only when BA was used at the concentration of 2.0 mg/l, and an inhibitive effect was observed with higher or lower concentration of BA. The number of regenerated buds reached up to 8.9 per explant with the combination of IAA 0.2 mg/l and BA 2.0 mg/l. Among cut modes, oblique cut performed the best for its effect on the number and quality of regenerated shoots and its convenience to manipulate. With the optimized hormone combination and oblique cut mode. citrus transformation with green fluorescent protein (GFP) gene was performed and twelve independently transformed plant lines were achieved. Southern blot hybridization confirmed the stable integration of GFP gene into the citrus genome. The successful transformation of this cultivar revealed that it is possible to introduce other genes with agronomic traits into it. Furthermore, these GFP expressing transgenic plants could serve as a visual marker material for citrus somatic fusion and sexual hybridization.