| Peony?Paeonia lactiflora Pall.?is a perennial herbaceous flower with the longest cultivation history in the genus Paeonia of the family Paeoniaceae.It has beautiful appearance,rich colors and wide application value.It has been loved by people over the years.Nowadays,the demand for peony in domestic and foreign markets is increasing.However,the current complete regeneration system of peony has not been realized and it is difficult to carry out factory production.Therefore,how to successfully expand the fine varieties of peony and realize large-scale production has become the primary problem to be solved.In this study,peony seed embryos were used as experimental materials to screen the best hormone combinations and culture methods for seed embryo initiation culture,cluster bud induction and proliferation,embryonic seedling rooting,callus induction and redifferentiation,and embryogenic callus.Solve the problems of low induction rate of cluster buds,difficulty in rooting,difficulty in redifferentiation of callus,difficulty in induction of embryogenic callus,etc.The regeneration system of peony was established,the optimal initiation culture mode of the seed embryo,the optimal culture mode of the callus,the optimal explants and hormones of the embryogenic callus were determined,which laid the foundation for the large-scale production.The main conclusions are as follows:1.The best sterilization method for seed embryos is 75%alcohol disinfection for 30 s,followed by 0.1%Hg Cl2for 5-8 minutes;the best embryo age suitable for germination of seed embryos is 75?95 d after pollination;the best culture method is The seed embryos were inoculated in the starting medium MS+0.5 mg·L-1GA3+1.0 mg·L-16-BA,cultured in the dark for 4 days,and then placed under normal light conditions for culture.The germination speed and growth status of the embryo seedlings Significantly improved.2.The two-step induction method can significantly increase the induction rate of cluster buds of peony embryos,grow vigorously,and reduce abnormal buds.The optimal medium for initial induction and secondary induction of cluster buds of seed embryos are MS+1.0mg·L-1GA3+1.0 mg·L-16-BA and MS+1.0 mg·L-1GA3+2.0 mg·L-16-BA,the induction rates were 48.33%and 63.89%respectively;the optimal medium for the proliferation of clump buds is MS+1.0 mg·L-1GA3+3.0 mg·L-16-BA+0.3 g·L-1CH+0.1 mg·L-1NAA+1.0 mg·L-1PVP,the average proliferation coefficient of cluster buds is 2.29,growing vigorously;the cluster buds of cotyledon nodes are the same as those of seed embryos The growth status under hormone culture conditions is quite different.The cotyledon node grows better in the initial induction,and the secondary induction effect of the seed embryo cluster bud is better than the cotyledon node cluster bud.3.The best rooting culture method of peony embryo seedlings is cultured in 1/2MS medium for 20 days,then transferred to 1/2MS+1.0 mg·L-1IBA medium for 35 days;1.0mg·L-1IBA and The superimposed use of 1.0 mg·L-1IAA can stimulate the occurrence of adventitious roots and increase their number,but callus will be produced around the radicle at this concentration.4.Cotyledons are the best explants for callus induction,dark culture for 20 days can significantly improve the quality of cotyledon callus induction;MS+0.5 mg·L-1TDZ+0.5mg·L-12,4-D+0.5 mg·L-1NAA is the best medium for callus induction;PVP is better than AC in improving browning,but only suitable for light browning;the most suitable medium for callus differentiation MS+0.5 mg·L-1TDZ+0.2 mg·L-1NAA+1.0 mg·L-1PVP,the differentiation rate was 10.71%,0.5 mg·L-1KT+0.2 mg·L-1NAA+1.0 mg·L-1PVP can also induce the callus to differentiate into adventitious buds,and the differentiation rate was5.71%.5.Peony cotyledons,hypocotyls and seed embryos failed to directly induce somatic embryos,but intact seed embryos can directly induce embryogenic callus,and 1.0mg·L-16-BA+1.0 mg·L-1NAA and 1.0 mg·L-16-BA+2.0 mg·L-12,4-D medium is better than adding 1.0 mg·L-16-BA+2.0 mg·L-1PIC,if PIC is used,its concentration needs to be increased appropriately;when embryogenic callus is induced through indirect route,1/2MS(Ca2+)+1.0mg·L-1TDZ+0.5 g·L-1CH+1.0 mg·L-1PVP+2.0 mg·L-1PIC is more suitable for ordinary callus to transform into embryogenic callus.At this concentration,the number of pale yellow raised grains is large,and embryogenic callus The induction rate is as high as 46.67%. |
PDF Full Text Request