Agrobacterium-mediated transformation of Japanese lawngrass (Zoysia japonica Steud.) containing a synthetic cryIA(b) gene from Bacillus thuringiensis.

摘要

Transgenic Japanese lawngrass plants containing a synthetic cryIA(b) gene from Bacillus thuringiensis under the control of a maize ubiquitin promoter were developed by Agrobacterium-mediated transformation. A total of 1540 embryogenic calluses derived from dormancy-removal mature seeds were co-cultured with the disarmed strain EHA105 harbouring the binary vector pKUB. Three days after co-culture with EHA105 in the dark at 21 degrees C, transient beta -glucuronidase (GUS) expression frequency was 74.2%. After selection with 100 mg/l hygromycin B, a total of over 50 independent resistant cell clones and 25 regenerated plants were obtained. The integration and expression of the cryIA(b) gene into the genome was confirmed in 22 regenerated plants by the GUS histochemical assay, PCR amplification, Southern blotting and Western blotting analysis, with a transformation efficiency of 1.4%. The entire process from callus induction of mature seeds to production of transgenic plantlets was 80-100 days. T1 progeny segregation analysis of these transgenic lines demonstrated that 59.1% of the transgenic events were inherited in a typical Mendelian fashion..