Pear Transformation With Polyphenol Oxidase Double-stranded RNA (dsRNAi)

The effects of various factors including different selecting period and material, basic medium ,concentration of phytohormone on adventitious bud induction,muitiplication and differentiation,rooting culture of Pyrus pyrifolia Nakai'Hosui' were studied.Result showed that the best tissue time culture period was in May to June and the optiuminduction medium was AS+ 6-BA 1.0 mg/L+NAA 0.2 mg/L+GA₃ 1.0 mg/L+Vc0.1 mg/L, witch was of benefit to adventitious bud muitiplication and browning decrease, the optimum rooting medium was ASH+0. 2mg/L NAA ,the Percentage of rooted shoots was 60%.Leaves which located in petiole and nervation after successive subculture for 28 to 35 days had high regeneration frequency.The time of prophase dark culture was 21 days and the light subculture was 50 days.The optimal medium for plant regeneration was NN69 supplemented with TDZ(2.0 mg/L) and IBA(0.1 mg/L), 86.7% regeneration rate and 2.92 buds per leaf were achieved. AgNO₃ enhanced the regeneration efficiency while its concentration varied from 0.1 mg/L to 1.5 mg/L ,and the most effective concentrations of AgNO₃ on treatment 12(NN69+TDZ2.0 mg/L.+IBA0.2 mg/L.) and treatment 15(NN69+TDZ1.5 mg/L+IBA0.3 mg/L) were 1.0 mg/L and 0.1 mg/L.The basal and middle leaf sections were more effective than apical leaf sections in inducing adventitious buds. Adventitious buds were easier induced with the leaf abaxial side touching media than with the leaf adaxial side touching media.Factors that influence percentage of transformation including time of dipping and co-culture, type and content of antibiotic, day of delayed selection culture, intension of selection were studied. The effective transformation system of Pyrus pyrifolia Nakai'Hosui'was developed as follow: Leaves were dipped into agrobacterium suspension for three minutes, then were co-cultured in darkness on NN69 medium with TDZ 2.0 mg/L, IBA 0.2 mg/L and AgNO3 1.0 mg/L for three days, and then cultured in darkness on NN69 medium with TDZ 2.0 mg/L, IBA 0.2 mg/L ,AgNO3 1.0 mg/L and Cef 250 mg/L. Selection culture was carried out also in darkness on NN69 medium with...