The Construction Of Project Agrobacterium Of Anti-herbicidal Bar Gene And Study Of Leaf Regeneration System In Strawberry

For exploring efficient method for strawberry(Fragaria ananassa Duch) anti-herbicidal bar Gene transformation by Agrobacterium tumefaciens the triparental mating hybridization was done to transform eukaryotic expression vector into Agrobacterium tumefaciens LBA4404.The products of mycelium PCR,plasmid PCR and plasmid digested by Xbaâ… and BamHâ… enzyme were tested by electrophoresis.The constructed project Agrobacterium was cultured with plantlet of strawberry.The strawberry cultivars Toyonoka suitably cultivated in Anhui Province were taken as material to establelish a simple and high-efficient leaf regeneration system.Callus and adventitious bud induction from leaf and rooting of in vitro shoot had been studied to provide a fundament for strawberry transgenic research.The main results gained were as follows.1,Utilizing triparental mating hybridization of LBA4404,HB101 and DH5αwas proved that the project agrobacterium with eukaryotic expression vector pBI121-bar had been constructed successfully by electrophoresis of products in PCR with bacterium colony and plasmid and in plasmid digested with Xbaâ… and BamHâ… enzyme.2,The critical concentration of kanamycin for mortality of untransgenic strawberry plantlet and callus from leaf was 25 mg/L and 20 mg/L respectively3,The critical concentration Cef and Carb efficient for control of project agrobacterium growth were both at 200 mg/L.4,The combination of 6-BA and NAA concentration cann't induce adventitious buds,they can only induce callus of strawberry leaf.5,The best culture medium for the adventitious bud induced from leaf was MS+6-BA 3.0 mg/L+2,4-D 0.2 mg/L+IBA 0.3 mg/L by analysis of SPSS software.6,The optimal rooting culture mediums for Toyonoka were 1/2MS+IBA 0.1 mg/L.7,The critical concentration Carb efficient for control of Agrobacterium rhizogenes R1601 growth was at 500 mg/L.8,When the OD₆₀₀ of Agrobacterium rhizogenes R1601 is 0.6,it is the most effect on frequency of transformation...