以万寿菊(Tagetes erecta)里程碑·黄色的小叶为外植体, 采用农杆菌介导法探究抗生素浓度、菌株类型、菌液浓度、侵染时间、共培养时间、乙酰丁香酮浓度和抗褐化剂种类对万寿菊遗传转化效率的影响。结果表明, 头孢霉素(Cef)和硫酸卡那霉素(Kan)的适宜浓度分别为100 mg·L^-1和10 mg·L^-1; EHA105菌株的稳定转化效率最高; 菌液浓度OD₆₀₀=0.1、侵染5分钟及共培养1天为最佳侵染条件。此外, 在侵染过程中添加100 µmol·L^-1乙酰丁香酮(AS)和筛选培养基中添加0.2 g·L^-1聚乙烯吡咯烷酮(PVP)均能提高出芽率。经GUS染色、PCR检测以及Southern blot检测证明转化成功, 转化效率达到4%左右。研究结果为万寿菊基因功能研究和转基因育种奠定了基础。

In this study, we used the leaflets of marigold (Tagetes erecta) Milestone Yellow as explants to investigate the major factors impacting the transformation efficiency of Agrobacterium-mediated method. The factors included antibiotic concentration, strain type, bacterial concentration, infection time, co-culture time, acetosyringone concentration and anti-browning agent type. We found that the suitable concentrations of cefotaxim sodium salt and kanamycin sulfate were 100 mg·L^-1 and 10 mg·L^-1, respectively. We also found that the strain EHA105 had the highest transformation efficiency up to 4%. The best infection conditions for EHA105 were at bacterial concentration of 0.1 for OD₆₀₀, infected for 5 minutes, and co-cultured for 1 day. In addition, the bud germination rate could be improved by both applying 100 μmol·L^-1 acetosyringone during the infection process and adding 0.2 g·L^-1 polyvinyl pyrrolidone in the screening medium. This study laid a foundation for marigold gene function research and transgenic breeding.