Production of transgenic plants of the Liliaceous ornamental plantAgapanthus praecox ssp orientalis (Leighton) Leighton viaAgrobacterium-mediated transformation of embryogenic calli

摘要

A system for producing transgenic plants was developed for the Liliaceous ornamental Agapanthus praecox ssp. orientalis (Leighton) Leighton via Agrobacterium-mediated genetic transformation. Leaf-derived embryogenic calli were inoculated with A. tumefaciens strain EHA101/pIG121Hm or LBA4404/pTOK233, both of which harbored the binary vector carrying the neomycin phosphotransferase II(NPTII), hygromycin phosphotransferase (HPT) and intron-containing beta -glucuronidase (GUS-intron) genes in the T-DNA region. Following co-cultivation, the calli were transferred to a medium containing 1 mg 1(-1) picloram (PIC), 50 mg 1-1 hygromycin and 500 mg 1(-1) cefotaxime, on which several hygromycin-resistant (Hyg(r)) cell clusters were obtained 5-6 weeks after transfer. Agrobacterium strain, co-cultivation period and acetosyringone (AS) treatment during co-cultivation affected the number of Hyg(r) callus lines produced: the best result was obtained when embryogenic calli were co-cultivated with LBA4404/pTOK233 for 7 days in the presence of 20 mg 1(-1) AS. Hyg(r) calli were transferred to the same medium, but lacking PIG, for inducing somatic embryos. Somatic embryos thus obtained developed into complete plantlets following their transfer to a medium without PIC and antibiotics. All of them were verified to be stable transformants by GUS histochemical assay, PCR and Southern blot analyses.