根癌农杆菌介导的高赖氨酸蛋白基因转化叶用莴苣的研究

TRANSFORMATION OF LETTUCE USING AGROBACTERIUM TUMEFACIENS

构建了植物表达载体pLBI,该载体携带 35S启动子、高赖氨酸蛋白基因 (LRP)、NPTII基因和NOS终止子 .对影响根癌农杆菌 (AgrobacteriumtumefaciensL .)转化的多种因素进行探索后 ,建立了一种农杆菌介导的稳定的叶用莴苣遗传转化系统 .选择目前生产上大量推广应用的叶用莴苣品种 ,以其无菌苗子叶为外植体 ,经农杆菌LBA4 4 0 4 (含质粒pLBI)感染 ,在含 ρ(Kan) /mgL-1=10 0的芽分化培养基上筛选转化芽 ,转入含相同浓度抗生素的生根培养基上进行生根筛选 ,直至得到完整的再生植株 .PCR扩增及Southernblot的检测结果均表明 ,高赖氨酸蛋白基因已整合到叶用莴苣基因组中 .图 4表 4参

Plant binary expression vector pLBI containing lysine-rich protein gene, 35S promoter, NOS terminator and NPTII gene was constructed. An efficient lettuce(Lactuca Sativa L.)transgenic system was established through explorating the factors affecting the transformation. The cotyledons from the sterile seedlings were used as the explants and infected with A.tumefaciens LBA4404 containing pLBI. The transgenic plants of regenerants was indicated by plant growth on medium containing ρ(kanamycin)/mg L -1=100. PCR and Southern blot analysis demonstrated further that lysine-rich protein gene had been integrated into transformants. Fig 4, Tab 4, Ref 6