A Preliminary Study Of Disease Resistance Gene Into Potato And Its Expression

In this experiment,a Agrobacterium mediated potato genetic transformation system was established,and three disease resistance genes(i.e.antifungal protein,non-specific lipid transfer protein and cyclophilin-like protein) were transformed into the genome of three common potato cultivars,Jinying 8,Dongnong 303 and Zihua 851 via this transform system.Regenerated Kan resistant plants were acquired,which were further subjected to specific primer PCR and pathogen resistance investigation.The results were as follows:1.Potato stem sections from sterile tube-plants of Jinying 8,Dongnong 303 and Zihua 851 were used as explants.The media for Dongnong 303 calli induction and adventitious bud regeneration were MS+1 mg/L 6-BA+0.2 mg/L NAA+0.2 mg/L GA₃+0.05 mg/L KT,and MS+1 mg/L 6-BA+5 mg/L GA₃,respectively,wheras the media for calli induction and adventitious bud regeneration of Jinying 8 and Zihua 851 were MS+2 mg/L 6-BA+0.2 mg/L NAA and MS+1 mg/L 6-BA+5 mg/L GA₃ respectively.1/2 MS was taken as rooting medium.75 mg/L kanamycin was chosen as the selecting pressure for callus inducing and adventitious bud regeneration.50 mg/L kanamycin was for rooting.200 mg/L cefotaxime was chosen to prevent the Agrobacterium from growing on the medium for the three potato cultivars.2.The optimized potato Agrobacterium-Mediated genetic transformation conditions were as follows:pre-culture stem section explants 2 days for calli induction,Agrobacterium with 0.5OD value infected the explants 10 minutes,cultured time 3 days in dark,25℃and 50 umol/L acetosyringone was used to stimulate the infection of explants by Agrobacterium,2200 mg/L CaCl₂ was added to calli induction media for a high transformation efficiency.3.Five Dongnong 303 plants,three Zihua 851 plants and six Jinying 8 plants with antifungal protein gene transformed;four Dongnong 303 plants,five Zihua 851 plants and five Jinying 8 plants with non-specific lipid transfer protein gene transformed;six Dongnong 303 plants,three Zihua 851 plants and five Jinying 8 plants with cyclophilin-like protein gene transformed were obtained.All these plants were further verified by specific primer PCR of antifungal protein, non-specific lipid transfer protein and cyclophilin-like protein genes,which improved that these three genes were transformed into the three cultivars. 4.Preliminary investigation of resistance disease showed that disease resistance level appeared to be enhanced in some of the transgenic plants.