The Effect of Growth Regulators and Antibiotics on Eggplant Transformation

Abstract

A protocol with a high rate of transformation and regeneration of `Hibush' eggplant (Solanum melongena L.) has been developed. This protocol used leaves of in vitro-grown seedlings as a source of explants. The shoot regeneration culture medium contained 0.1 μm thidiazuron (TDZ) combined with 10 to 20 μm N6-[isopentyl] adenine (2iP). Adding TDZ significantly improved regeneration efficiency and produced a mean of 15 buds and 3 to 4 shoots per explant. When explants were cocultivated with Agrobacterium tumefaciens strains Q10, Q20, Q30, Q40, Q201, Q202, Q203, or Q204 containing the native cryIIIB Bacillus thuringiensis (Bt), neomycin phosphotransferase (NPTII), and β-glucuronidase (uidA) genes, a callus/bud regeneration frequency of 38.8% was observed on the selection medium. Kanamycin at 50 μg·mL-1 was most effective in selecting for transgenic buds and shoots. Augmentin at 300 μg·mL-1 was used to eliminate A. tumefaciens. Augmentin also enhanced shoot proliferation. A transformation/regeneration efficiency of 20.8% was observed for shoot production. More than 400 putative transgenic plants have been produced with this method. From 50 putative transgenic plants, gene integration has been confirmed with Southern blot analysis and progeny tests.