| In this article , factors influencing regeneration of maize immature embryos of inbreds J7-3, B73, B77 and hybrids J7-3 A188, B73 A188, B77 A188 have been studied. An high efficient regeneration system of inbreds J7-3 and B73 were developed. Also, factors affecting maize transformation mediated by Agrobacterium tumefaciens have been examined through GUS transient expression. GM-CSF gene transplants were abtained using the developed regeneration and transformation system. Main results are as following:1. In used culture conditions, genotype is the main factor determining regenerable ability of maize immature embryo . and hormone composition, days from formation of embryogenic callus to rededifferention medium, growth state of immature embryos are also affecting factors. It is better to use N6 medium for inducing and use MS medium for redediffirention than only use N6 medium or MS medium for regeneration. When inducing the medium should containing 1.0mg/L 2, 4-D, Dicamber and AgNO3. Embryogenic callus should be moved to regeneration medium after 6-8 days since they appeared. Higher frequence of normal plantlets can achieved through add a small quantity of ABA (0.03mg/L) to rededifferention medium. It may take only 40 days from immature embryo to plantlet formation.2. 9 minutes of infecting , 3 days of co-cultivation and 4-5 days of resting before selection is good for transformed cell to resume. In addition, 1.0- 1.5mm length of immature embryos is easy for transformation and regeneration after being infected. Infecting basic medium, osmotic pressure, glucose concentration et al. are also factors affecting transformation.3. The resistant plants of inbred B73 and hybrid B73 X A188 were gained through selection after being infected. The results of PCR and PCR-Southen analysis of transformants indicated that exogenes has been inserted into maize genome. |
