Study On Regeneration System And Genetic Transformation System Of Populus Hopeiensis

In this study,the 1-year-old shoots of Populus hopeiensis were used as explants to establish an efficient regeneration system of Populus hopeiensis.On the basis of this,the paper takes the non vaccine leaves of Populus hopeiensis as the acceptor material,uses the Agrobacterium mediated transformation of WOX gene,and studies the main factors affecting the transformation efficiency,which provides ahigh-efficiency,safe and large-scale genetic transformation and improvement platform for Populus hopeiensis,and provides theoretical and technical reference for other poplar genetic transformation and cultivation of new varieties,and functions of related genes the establishment of poplar genetic transformation system has important practical significance.The main results were as follows:1.Establishment of regeneration system of Populus hopeiensis:the best disinfection time was 30 s with 75%alcohol and 12 min with 0.1%HgCl2,the best disinfection effect,the lowest pollution rate was 10%at this time,and death rate was 13.33%;the best medium for inducing axillary bud of stem segment was 1/2MS+0.5 mg·L-1 6-BA+ 0.05 mg·L-1 NAA,the germination time of axillary bud was 7 days,the induction rate was 95%;the best rooting medium was 1/2MS+0.3 mg·L-1 IBA+0.05 mg·L-1 NAA,the rooting rate is as high as 100%;The results showed that the mixture of peat soil,vermiculite and perlite at 6:3:1(v/v/v),The survival rate was 93.7%.2.Establishment of genetic transformation receptor system of Populus hopeiensis:the best medium for inducing adventitious buds was MS+0.3 mg·L-1 TDZ+0.4 mg·L-1 IBA.After dark culture for 12 days,the leaves are transferred to low light for cultivation,which is conducive to the differentiation of adventitious buds.The average number of adventitious buds was 11.04;the selected pressure of Hyg B for inducing adventitious buds and rooting was 3.0 mg·L-1,and will not affect the leaves Adventitious buds and rooting were induced.The concentration of temetin was 400 mg·L-1,which can effectively inhibit the growth of Agrobacterium,and has the least effect on the differentiation of leaves.3.Optimization of genetic transformation conditions of Populus hopeiensis:the best transformation conditions of Agrobacterium tumefaciens mediated by Populus hopeiensis were as follows:1 day of pre-culture,10 minutes of infection in the bacterial solution with OD600=0.6,3 days of co-culture,58 resistant plants were obtained through selective culture of adventitious bud regeneration and rooting.PCR detection of 58 resistant plants showed that 17 of them were positive,and the transformation rate was 29.3%.which indicated that WOX gene had been introduced into the genome of Populus hopeiensis.