摘要
以油菜子叶节为外植体优化了农杆菌介导的甘蓝型油菜的遗传转化体系,为在油料作物中用基因工程手段生产γ-亚麻酸打下基础。采用CaCl2法制备农杆菌AGL1感受态细胞,将已经构建好的植物表达载体pPZP221(35S-D6D-NOS)通过液氮冻融法转化农杆菌,PCR筛选阳性转化子。结果表明:优化的甘蓝型油菜子叶节外植体预培养条件是:MS+2.0 mg/L 6-BA+0.2 mg/L NAA培养基预培养2 d;用于侵染的农杆菌菌液的最适浓度为OD600=0.4,时间为5 min,并得到转基因植株。经侵染的外植体在含有庆大霉素的分化选择培养基上培养,抗性芽移至生根培养基,PCR检测阳性小植株移栽花盆,共得到12株转基因植株,首次将来自月见草的D6D基因通过农杆菌介导法整合到油菜基因组中。
The Δ6-fatty acid desaturase gene from Oenothera biennis was introduced into the explants of Brassica napus in this study,and postive transgenic plantlets were confirmed with PCR which contribute to provide the primary basis for the future application of gene engineering to γ-linolenic production.This study used cotylendon node explant to optimize Agrobacterium-mediated Brassica napus genetic transformation system.The optimal pre-culture condition for the cotylendon node is two days on MS medium＋6-BA 2.0 mg/L＋NAA 0.2 mg/L.Agrobacterium was transformed with binary vector pPZP221 harboring 35S promoter-driven ObD6D gene,which was used to infect brassica napus cotylendon node with optimal concentration OD600=0.4 and time 5 min.Infected explants were cultured on differential selective medium with gentamicin and the resisitant explants were subsequently transfered to root-promoting medium.Postive transgenic plantlets were detected by PCR and transfered to soil.In total,we got twelve transgenic plantets.
出处
《华北农学报》
CSCD
北大核心
2011年第6期50-54,共5页
Acta Agriculturae Boreali-Sinica
基金
教育部“春晖计划”科研项目(Z2004-2-15027)
关键词
油菜
月见草
Δ6-脂肪酸脱氢酶基因
Γ-亚麻酸
农杆菌介导
Brassica napus
Oenothera biennis
Δ6-fatty acid desaturase gene
γ-linolenic acid
Agrobacterium-mediated
