【摘要】 以白三叶萌动种子作为转化受体,利用npt-Ⅱ筛选基因对影响农杆菌介导的遗传转化外部因素进行深入探索,通过优化转化条件,建立了农杆菌浸种途径介导的白三叶遗传转化体系。白三叶种子催芽1 h后,加入OD600值为0.9的农杆菌菌液,附加5 min+30 s真空处理方式,振荡侵染24 h,共培养1 d后,用50 mg/L卡那霉素筛选抗性植株,该体系能够获得较好的转化效果,阳性转化率为45.5%。通过PCR检测和GUS组织化学染色法,初步验证npt-Ⅱ和GUS基因已经整合到植物基因组中。更多还原

【Abstract】 The germinating seeds of white clover were used as transformation receptor,and npt-Ⅱgene was chosen as a selection gene.External factors that affect the efficiency of genetic transformation were studied and optimized.A high-frequency transformation system of white clover had been established by adopting Agrobacterium tumefaciens soaked seeds transformation technology.The seeds of white clover germinated for 1 hour,then adding Agrobacterium with concentration of 0.9(OD600) under 5 min+30 s vacuum treatment.After 24 hours infection in oscillating culture and 1 day co-cultivation,the transferred plants were screened by 50 mg/L kanamycin.This transformation system was so effective,that positive transformation rate reached 45.5%.The transgenic plants were also examined by PCR analysis and GUS staining.It was proved that npt-Ⅱand GUS gene had been integrated into the genome.更多还原