Establishment And Optimization Of Regeneration And Genetic Transformation System Of Apple Rootstock G.41

Apple rootstock plays a very important role in production.Its performance on scion traits,such as tree potential of the scion,fruit size,fruit maturity and nutritional value also plays a very critical role.This study established regeneration system of apple rootstock G.41 by the method of tissue culture,at the same time genetic transformation system of the G.41 also was established and optimized.The plant expression vector of MpNCED2 genes was successfully constructed using Gateway carrier construction technology,and it was transferred into the plant of Arabidopsis and apple rootstock G.41 by Agrobacterium-mediated method,both of the resistance transgenic plants were obtained,respectively.The study laid foundation for the application of G.41.The main test results are as follows:1.A simple and efficient protocol for obtaining shoots from leaf explants was established by optimizing the combinations of plant growth regulators,mode of wounding,and explant orientation on the culture medium.The best shoot multiplication index(2.58)was obtained from successful subculture medium that was the standard MS medium supplemented with 3.55 ?M BA,0.16 ?M IBA.Regeneration rates were highest(99%)when MS medium was supplemented with 2.7 ?M TDZ and 0.9 ?M NAA,and cut-wounding explants before placing the abaxial surface in contact with the medium.The best rooting percentage(80%)was obtained on MS medium supplemented with 4.92?M IBA.Plantlets were rooted in vitro and survived acclimatization in the laboratory and greenhouse.2.Through a genetic transformation system of the apple rootstock G.41 was optimized and established,we got the optimum Kana selection pressure of leaf was 4 mg/L and the optimal Cef concentration of leaf was 200 mg/L.3.Using Gateway carrier construction technology successfully constructed the plant expression vector of MpNCED2 genes,and it was transferred into the plant of Arabidopsis by Agrobacterium-mediated method,then 11 T3 overexpression transgenic Arabidopsis plant were acquired.At the same time the plant expression vector was transferred into apple stock G.41,and the resistance of the plants also was achieved.In the process of transformation,suitable drying time for helping to agrobacterium infect blades was 5 min;Silwetl-77 concentration for advantageous to the agrobacterium infect blades was zero.