| Dormant buds taken from one year old persimmon branches,as in vitro explants materials,to study the early generations, subculture, leaf callus and buds induction. The paper laid the foundation of study the dwarfing mechanism from molecular aspects and physiological traits aspects.At the same time promote the rapid breeding and production promotion of this dwarf varieties of persimmon. Results as following:1. Use 10%H202 on dormant buds for disinfection 10min and 20min, the survival rate was 68% and 80%,Disinfection effect is obviously better than use 5%NaC10 and 0.1%HgCl. Reactive oxygen species can promote dormant buds germinate which produced by the decomposition of hydrogen peroxide. Solved the problem of dormant buds easy browing and death.After comparing the three basic medium found that the tissue plants grow in Modified MS midium (Murashige and Skoog medium with KNO3 and NH4NO3 reduced to half of the original strength) can up to 15.9mm, suitable for persimmon early generations.The results of orthogonal design trial show that dormant buds in (1/2N)MS medium added 0.05mg/LNAA+1mg/LZT can grow very well with small callus.2. Seedlings that subcultured in vitro for 4-5 generations, were cultured in media of (1/2N) MS+ZT1.0mg/L+IAA0.05mg/L+GA30.05mg/L+PVP500mg/L+Sucrose 30 g/L+ Agar 6.0g/L and (1/2N)MS+ZT1.0mg/L+IAA 0.05 mg/L+PVP 500 mg/L+Sucrose 30g/L+Agar 6.0g/L by turns. Axillary buds of the previous generation were cut and inserted into the appropriate media for propagation, with each medium for 25-30 days. Using this procedure can effectively solve the problem that Diospruskaki Linn. cv. Nantongxiaofangshi couldn’t be propagated in vitro with lack of clump seedlings.By studying the growing situation, plant height and propagation coefficient of Nantongxiaofangshi impacted by different concentrations of ZT, BA and TDZ, we found that the frequency of axillary buds appearing from seedlings cultured in media of lmg/LZT was significantly higher than that in any other treatment Which indicated the optimum culture condition for Nantongxiaofangshi.The comparison of growing situation, plant height and propagation coefficient of Nantongxiaofangshi impacted by different concentrations of IAA, IBA and NAA revealed that low concentration of auxin especially IAA and IBA in 0.05mg/L can stimulate the elongated growth and sprouting of axillary bud of Nantongxiaofangshi cultured in vitro, which is propitious to axillary bud propagation.3. Optimize the basic culture medium affecting the differentiation of leaf regeneration,TDZ, NAA, and the days of dark cultivation by L9 (34) orthogonal experiment design.The best processing combination obtained by orthogonal experiment analysis is 3 mg/L TDZ+0.2 mg/LNAA+1000 mg/L PVP+(1/2 N) MS medium+14 days dark cultivation and then thirty days later transferring them into MS (1/2)+ZT 2.0 mg/L+ IAAO.l mg/L medium to induce the growth of adventitious bud.This kind of medium induce the formation of callus reach up to 82.52% and the average number of adventitious bud as much as 0.92 and the differentiation rate is 56.31%, which is suitable for callus induction and adventitious bud regeneration of leaves of Nantongxiaofangshi.Medium added different concentration of NAA,IBA,IAA,2,4-D and TDZ,comparing the different kinds of auxins affecting differentiation rate of callus and adventitious bud numbers and differentiation rate on cells of small square persimmon leaves.NAA affecting on the formation of the blade callus differentiation is better than other kinds of auxins.0.2 mg/L NAA inducing the number of adventitious bud up to 0.57 in the whole processing,which is the most.And the differentiation rate up to 36.07%, significantly higher than other processing of average number of adventitious bud and differentiation rate. |
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