摘要： 为获得抗旱性较强的转基因苜蓿植株,以紫花苜蓿品种中苜2号作为基因转化受体,通过对外植体选择、菌液浓度、选择压确定、侵染时间和共培养时间等因素进行优化,成功建立了农杆菌介导的遗传转化体系。在此基础上,将沙冬青脱水素基因通过农杆菌的介导,转化到中苜2号中。试验结果显示,子叶和下胚轴作为外植体愈伤组织诱导频率最高,可达100%;获得抗性愈伤组织与转基因植株的卡那霉素最佳筛选浓度为25 mg/L;外植体与农杆菌的共培养时间以5 d为最佳。试验共获得126株T0抗性株,PCR检测30株表现为阳性,表明脱水素基因已转入受体植株中。
关键词:
紫花苜蓿,
遗传转化,
脱水素基因(AmDHN)
Abstract: In order to obtain transgenic plants of resistant drought，by comparative analysis on the different explant tissues，concentrations of Agrobacterium and kanamycin，time of inoculation and coculture，etc，an Agrobacterium-mediated transformation for an alfalfa variety Zhongmu No. 2 was established. On the basis of the transformation，?dehydrin of Ammopiptanthus mongolicus was transformated into genome of Zhongmu No. 2 by A. tumefaciens-mediated. The results showed that cotyledon and hypocotyl was the best explant for callus induction and transformation.?The optimal kanamycin concentration for selection of resistant callus and transgenic plant was 25 mg/L in the selection medium. The explants and bacteria was cocultured for 5 d. One hundred and twenty-six independent resistant?plants were obtained，30 plants were confirmed to be transgenic by PCR amplification of AmDHN gene and 35S?gene.
Key words:
Alfalfa,
Genetic transformation,
Dehydrin
中图分类号:
聂利珍, 郭九峰, 孙杰, 孙国琴, 刘永志. 沙冬青脱水素基因转化紫花苜蓿的研究[J]. 华北农学报, 2012, 27(3): 96-101. doi: 10.3969/j.issn.1000-7091.2012.03.019.
NIE Li-zhen, GUO Jiu-feng, SUN Jie, SUN Guo-qin, LIU Yong-zhi. Research on Transformation of DHN Gene from Ammopiptanthus mongolicus into Alfalfa(Medicago sativa L. )[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2012, 27(3): 96-101. doi: 10.3969/j.issn.1000-7091.2012.03.019.