摘要
[目的]为利用转基因技术培育中华补血草新品种奠定基础。[方法]对农杆菌介导的中华补血草转化体系中的各种影响因素进行了系统研究,以构建中华补血草的遗传转化体系。[结果]中华补血草叶片对卡那霉素敏感,以50 mg/L卡那霉素作为补血草叶片转化受体较为适宜。400 mg/L特美汀能有效抑制农杆菌,而500 mg/L头孢曲松钠则不能。菌株EHA105对中华补血草的转化率达15%,远高于菌株LBA4404和GV3101。对于侵染中华补血草无菌苗叶片的重悬菌液,以OD600为0.7-1.0为宜。4 d共培养的效果较好,15.6%外植体可分化成芽。PCR检测结果表明30株卡那霉素抗性植株均为阳性,而阴性对照未扩增到条带,表明NPTⅡ外源基因已整合到受体植物基因组中。[结论]影响中华补血草转化率的关键因素是农杆菌菌株的选择。
[Objective] The research aimed to lay the foundation for breeding new varieties of Limonium sinense by using transgenic technologies.[Method] The influencing factors in the transformation system of L.sinense mediated by Agrobacterium tumefaciens were systemically studied to construct the genetic transformation system of L.sinense.[Result] The leaves of L.sinense were sensitive to kanamycin and it was suitable to take 50 mg/L kanamycin as the transformation acceptor of L.sinense leaves.400 mg/L timentin could inhibit A.tumefaciens effectively,but not 500 mg/L ceftriaxone sodium.The transformation rate of Strain EHA105 to L.sinense reached 15%, greatly higher than that of Strain LBA4404 and Strain GV3101.For the suspended bacteria liquid that infected the leaves of L.sinense aseptic seedlings,its suitable OD600 was 0.7～1.0.The effect of co-culture for 4 d was better and 15.6% explants could be differentiated into buds.The results of PCR detection showed that 30 plants with the resistance to kanamycin were all positive and no band was amplified in negative control,which indicated that NPTⅡ exogenous gene had been integrated into the genome of acceptor plant L.sinense.[Conclusion] The key factor that affected the transformation rate of L.sinense was the strain selection of A.tumefaciens.
出处
《安徽农业科学》
CAS
北大核心
2008年第7期2668-2669,共2页
Journal of Anhui Agricultural Sciences
基金
国家重点基础研究发展"973"计划(2006CB100100)
关键词
中华补血草
农杆菌
遗传转化
影响因子
PCR
Limonium sinense
Agrobacterium tumefaciens
Genetic transformation
Affecting factors
PCR
