Study On Senescence Retarding Through P_SAG12-ipt Introduction Into Zoysia Sinica Hance

Zoysia sinica Hance was used in this study. the purpose of the study was to show a postponement of leaf senescence and to a prolonging green period of leaves by transferring P_SAG12-ipt gene into wild Zoysia. The results as follows:1. An efficient regenerating system of Zoysia was established. The inducing rate of calli reached 34.2% and 40. 8% respectively in modified MS culture medium added 2mg/L 2,4-D or 3mg/L Dicamba, using mature embryo as explant. 6-BA with low concentration was vital for transformation from non-embryogenic calli to embryogenic calli on subculture medium. The optimum condition for embryogenic calli inducing was obtained. Modified MS added 0.1mg/L 6-BA and 2mg/L 2,4-D or 3mg/L dicamba was best subculture medium, while modified MS added 0.2mg/L dicamba and 0. lmg/L 6-BA for differentiation, in which the regeneration rate reached 51. 8%. Modified MS with 0.2mg/L 2,4-D and 0.1mg/L 6-BA could be used as differentiation medium, but the regeneration rate was lower as 38. 6%.2. ipt gene controlled by the promoter of SAG12 was introduced into embryogenic callus of Zoysia by particle bombardment. 37 transformants were obtained after successively G418 resistant selection, PCR analysis and Southern blot. Among those plants, only 6 lines were detected as leaf senescence delayed and green period prolonged. Furthermore, the contents of CTK, soluble protein of transgenic plants were higher than that of wild plants, but ABA content of transformants was lower. The delayed senescence of transgenic plants was also proved by slowed reducement of photosynthetic rate of detached leaves.3. P_SAG12-ipt was transformed into the embryogenic calli of zoysia by Agrobacterium-mediated method successfully with NTPII as resistant gene. 35 transformants were achieved after selection in medium with 100mg/L G418. The result of PCR and Southern blot showed that P_SAG12-ipt gene had been transferred and integrated into Zoysia genomes. Among those transformants, the content of CTKs was enhanced evidently during the late growth period, the degradation of chlorophyll was retarded, the yellow turning date was postponed for about 15 days and the senescence process was inhibited in 5 transgenic Zoysia lines.4. Shadow and low-temperature are the major stress factors for Zoysia. The impact of shadow and low-temperature on the induction and expression of P_SAG12-ipt gene in transgenic Zoysia was investigated. The P_SAG12-ipt expression showed triggered and up-regulated by shadow and low-temperature, and the induced senescence was retarded.