农杆菌介导的燕麦创伤胚转化体系研究  

Study on Transformation System of Oat Trauma Embryo with Agrobacterium Mediated Method

作　　者：张丽君[1,2] 刘龙龙[2] 孙毅[3] 张建珍[1] 崔林[2] 

机构地区：[1]山西大学应用生物学研究所,山西太原030031 [2]山西省农业科学院农作物品种资源研究所农业部黄土高原作物基因资源与种质创制重点实验室,山西太原030031 [3]山西省农业科学院生物技术研究中心,山西太原030031

出　　处：《河北农业科学》2015年第3期49-54,76,共7页Journal of Hebei Agricultural Sciences

基　　金：国家燕麦荞麦产业技术体系项目(CARS-08-A-1);山西省自然科学基金项目(2014011030-1)

摘　　要：通过建立遗传转化体系,培育具有特定功能的转基因作物品种对改善作物的生存能力和生态环境适应力具有重要意义。参照农杆菌划胚介导植物萌发种子基因转化方法,以农杆菌菌株LBA4404(含质粒p CAMBAR.CH I.11)侵染转化燕麦种子,对转化体系中的菌液浓度、超声波功率、种子处理方式和抗除草剂筛选等不同转化条件分别进行了研究,以探索农杆菌介导的燕麦最适宜的遗传转化体系。结果表明:燕麦种子浸泡4 h后采用穿刺法划伤种子,1 000 W超声波处理划伤种子10 min,在OD600=0.4的农杆菌溶液中共培养2 d,最有利于种子转化;用0.8 mg/L草丁膦溶液与种子共培养能够有效抑制种子萌发进行初筛,用1 mg/L草丁膦溶液喷施幼苗进行除草剂抗性筛选,能够明显抑制假阳性苗的生长;将来自链霉菌(Streptomyces hygroscopicus)的Bar基因导入燕麦,经除草剂筛选以及Bar基因的PCR检测和Southern杂交分析证明获得转基因植株。Cultivating transgenic crop varieties with specific function through the establishment of genetic transformation system is significant for the improvement of crop survival ability and ecological environment adaptability.Referring to the gene conversion method of seed germination with agrobacterium mediated , the oat seeds was infected and transformed with agrobacterium strain LBA4404 （ contain plasmid pCAMBAR.CH I.11 ） , the transformation conditions including concentration of bacteria, ultrasonic power, processing method of seeds, selection of herbicide resistance and so on in the transformation system were studied to explore the most appropriate genetic transformation system of oats mediated by agrobacterium.The results showed that after immersed for four hours, oat seeds were scratched by puncture method, 1 000 W ultrasonic treatment by 10 min was done for the scratched seeds, then they were cultured in agrobacterium liquid concentration OD600 =0.4 for 2 days, it was the best condition for genetic transformation.Seeds cultured with 0.8 mg/L glufosinate concentration could effectively inhibit germination of oat seeds.Seedlings were sprayed by 1 mg/L glufosinate concentration to select herbicide , it could obviously inhibit the growth of false positive seedlings .The Bar gene was introduced into the oat , and transgenic plants were first screened by Basta then detected by PCR and Southern blot approaches.

关 键 词：燕麦 转化体系 BAR基因 农杆菌介导 

分 类 号：S512.603[农业科学—作物学]