根癌农杆菌介导的人骨形成蛋白-3成熟肽基因向油菜转化的研究

用根癌农杆菌(Agrobacteriumtumefaciens)共培养法,将骨形成蛋白-3成熟肽(hBMP-3m)基因导入甘蓝型自交系油菜(BrassicanapusL.)品种秦油-3号,并获得了hBMP-3m基因整合到油菜基因组中的植株。试验所用外植体为油菜4～5d苗龄的下胚轴;根癌农杆菌为LBA4404,其Ti质粒pCAMBIA-hBMP3m含hBMP-3m基因。切取2mm左右的下胚轴侵染根癌农杆菌后在分化培养基(MS+5.0mg/L6-BA+0.3mg/LNAA)上共培养2d,然后转到附加25mg/L潮霉素和500mg/L羧苄青霉素的分化培养基上。切下分化出的茎芽,插入附加25mg/L潮霉素的生根培养基(MS+0.4mg/LIBA)中,形成完整的植株。经PCR检测和Southernblot分析,证实hBMP-3m基因已插入到油菜细胞基因组中。

Transformation of Brassica napus with human bone morphogenetic protein 3 mature piptide gene mediated by Agrobacterium tumefaciens

Seeds of Brassica napus L.cv."Qinyou-3"were surface-sterilized and germinated hormone-free MS medium.After four to five days,the hypocotyls were excised in such a way.These hypocotyls were used as the explants for tissue culture and genetic transformation.After 2 days of co-cultivation with (Agrobacterium) tumefaciens strain LBA4404,the plasmid of which contains gene of hBMP-3m,the explants were transferred onto selection medium containing 25 mg/L hygromycin and 500 mg/L carbenicillin.About one month,shoots emerged from the explants inoculated.They were excised and transferred onto rooting medium containing 25 mg/L hygromycin.Whole plants were obtained.Transformation was confirmed by PCR and Southern blot analysis.