Research On Regeneration System And Agrobacterium Genetic Transformation System Of Banana Brazil(Musa Spp.)

The experiments were conducted on Brazil banana regeneration system and Agrobacterium-mediated transformation system. Regeneration system include regeneration system of the banana stem microdisk meristem, regeneration system of stem callus, regeneration system of male flowers embryonic callus. Microdisk stem slices were used for Agrobacterium-mediated transformation as explants. Optimization of the bacterial concentration included infection time, co-culture time, co-culture temperature and antibacterial agents. The main research results as follows:1. Microdisk stem slices efficiently regenerated in the MS+0.5mg/LNAA+4mg/L6-BA medium and in the dark at26℃.2. The different varieties of stem incubated in the MS+2,4-D0.7mg/L+6-BA4mg/L medium and in the dark at26℃, and the maximal inducing rate of callus was75%. However, only Banana Brazil (blue bar) and Hainan Dwarf banana can induce embryogenic callus, induction rate was0.1%and0.5%.3. The immature male flowers were cultured in the MS+2,4-D4mg/L+IAA1mg/L+NAA1mg/L medium and in the dark at26℃. The induction rate of embryogenic callus was3-4%.4. Agrobacterium-mediated transformation was researched by using microdisk stem slices as explant. The best transient expression of GUS was obtained under the conditions as follows:the Agrobacterium bacterial concentration was0.8(OD600) and the samples were infected for8min and co-cultured for4days at26℃in the dark.5. Timentin (300mg/L) was an appropriate antimicrobial agents for inhibiting agrobacterium, and the appropriate concentration of PPT was0.1-0.2mg/L for choosing.6. The Agrobacterium-mediated genetic transformation system of microdisk stem slice has been established from low-generation test-tube seedling in M. paradisiaca. Using the system, the GUS gene was transformed to banana (Musa paradisiaca) microdisk stem slice and transgenic seedlings were obtained. The PCR assays proved the target gene was integrated into the plant genome. The histochemical assays revealed that the GUS enzyme was steadily expressed in the transgenic plants.