摘要
系统研究了影响根癌农杆菌介导的萝卜（Raphanus sativus L.）遗传转化的若干因素,建立了萝卜遗传转化体系：即切取萝卜带柄子叶外植体,先进行2 d预培养,用OD600为0.3-0.5的EHA105农杆菌菌液侵染5-7 min后,再进行5 d共培养,然后将外植体转接到MS＋6-BA 6 mg.L-1＋NAA 0.05 mg.L-1＋Cef 500 mg.L-1的培养基上进行7 d延迟筛选,再将外植体转接到MS＋6-BA 6 mg.L-1＋NAA 0.05 mg.L-1＋Cef 500 mg.L-1＋Hyg 5 mg.L-1的培养基上进行10 d的抗性筛选。结果共获得126个抗性芽,其中8个抗性芽经PCR检测扩增出目的基因的启动子BcA9,约850bp,阳性率为6.3%,初步验证目的基因已经转入萝卜再生芽中。
The factors for Agrobacterium tumefaciens-mediated genetic transformation in Radish（Raphanus sativus L.）were studied.A radish genetic transformation system is established.Cotyledons with petiole were cultured in MS ＋ 6-BA 6 mg·L-1 ＋ NAA 0.05 mg·L-1 for 2 days as pre-culture.They were then infected for 5-7 min by the EHA105 Agrobacterium at OD600 = 0.3-0.5,and co-cultured in same medium for 5 days.The explants were then transferred to MS ＋ 6-BA 6 mg·L-1 ＋ NAA 0.05 mg·L-1 ＋ 500 mg·L-1 Cef medium for 7 days for pre-screening.The explants were later cultured no MS ＋ 6-BA 6 mg·L-1 ＋ NAA 0.05 mg·L-1 ＋ 500 mg·L-1 Cef ＋ 5 mg·L-1 Hyg medium for 10 days for selecting transgenic events.We got 126 resistant shoots,and 8 of them were amplified the target gene BcA9 with size about 850 bp.The preliminary validation showed that the target gene has been transferred into radish.
出处
《中国瓜菜》
CAS
2011年第4期18-21,共4页
China Cucurbits And Vegetables
基金
国家科技支撑计划项目(2009BADB8B03-1)
关键词
萝卜
遗传转化
细胞质雄性不育基因
Radish
Genetic transformation
Cytoplasmic male sterility gene
