Construction Of Efficient In Vitro Regeneration System Of Potato And Optimization Of Genetic Transformation Of Microtuber

Potatoes are both food and vegetable plants with large market demand.However,the potato industry is still facing the problems of low breeding level,insufficient supply of high-quality seed potatoes,the level of industrialized production still needs to be improved.Therefore,in vitro regeneration and genetic transformation of potato were studied in order to lay a foundation for the cultivation of new varieties and large-scale production of potatoes.In this study,potato cultivar ‘Atlantic',‘Kexin 23',‘Youjin' were used as material to establish and optimize in vitro regeneration system of stem segment,leaf,microtuber,field potato buds,field potato tuber.The genetic transformation and application of the potato variety ‘Atlantic'in vitro was studied using potato slices as receptor system.In the process of in vitro regeneration system construction,the effects of different plant hormone concentrations and proportions,different disinfection methods on the regeneration process of different explants were studied.The results are as follows.(1)The optimum medium for the adventitious buds induction from different explants was MS + 3 mg/L 6-BA + 0.2 mg/L NAA + 1.5 mg/L ZT(direct induction of stem segments),MS + 2 mg/L 6-BA + 0.6 mg/L NAA + 2 mg/L ZT(indirect induction of stem segments),MS + 3 mg/L 6-BA + 0.2 mg/L NAA + 2 mg/L GA3(direct induction of leaves),MS + 2 mg/L 6-BA + 0.5 mg/L NAA + 3 mg/L GA3(indirect induction of leaves),MS + 2 mg/L ZT + 1.5 mg/L IAA + 1 mg/L 6-BA(microtuber),MS + 2 mg/L 6-BA + 0.8 mg/L NAA(field potato bud),MS + 3 mg/L 6-BA + 2.5 mg/L NAA(field potato tuber);(2)The suitable medium for adventitious bud elongation was MS + 1.5 mg/L GA3 + 1.5 mg/L ZT + 0.5 mg/L NAA,the elongation efficiency was about 93.47% and the average shoot length was 2.46 cm;(3)The best method of adventitious root induction was MS + 0.01 mg/L EBR + 0.1 mg/L NAA,its rooting frequency was approaching 100%;(4)Field potato buds were soaked in 0.1% mercury for 8 minutes,then in 10% sodium hypochlorite solution for 3 minutes,the sterility rate was 95.27%;field potato tuber was soaked in 0.1% mercury for 13 minutes,the sterility rate was 75.98%.Using the slices of ‘Atlantic' microtuber as foreign genes transferring target,the effects of some basic parameters mediated by Agrobacterium tumefaciens on potato genetic transformation system were studied.A preliminary genetic transformation system for in vitro potato slices was established.The results showed that the optimal concentration of acetosyringone(50 ?mol/L),bacteria concentration(OD600=0.5),time of elution(5 min),co-culture time(3 d)could effectively improve the transformation frequency of the genetic transformation system of potato in vitro slices.The transgenic resistant potato seedlings were successfully obtained by using this transformation system.The yield of resistant buds was about 43.33% and the transformation efficiency was about 32.67%.