84 K杨再生和遗传转化体系的优化

为了优化84 K杨的再生和遗传转化体系,以84 K杨树叶片为外植体,采用正交试验研究了不同培养基和激素配比对芽和芽苗生根的影响,并从预培养时间、浸染菌液浓度、浸染时间、共培养时间及添加AS(乙酰丁香酮)5个方面,以卡那霉素抗性苗频率为衡量指标,研究了各因素对84 K杨遗传转化率的影响。结果表明,84 K杨叶片最适的芽诱导培养基及激素配比为MS+6-BA 0.5 mg/L+NAA 0.05 mg/L,芽苗最适诱导生根培养基及激素配比为1/2 MS+NAA 0.05 mg/L+IBA 0.05 mg/L;优化筛选的最适转化体系为:农杆菌活化菌液用MS液体培养基稀释10倍,浸染5 min,共培养4 d。采用上述优化的再生和遗传转化体系,84 K杨每片叶平均生芽数可达20个,芽苗生根率可达99%,生根数为6条,叶盘转化率可达35%。

Prioritization of the system of regeneration and genetic transformation of Populus 84 K

To prioritize the system of regeneration and genetic transformation of Populus 84 K,influences of different medium and proportion of hormone on bud and root of leaf-explant were studied by orthogonal designing.The impact of five aspects(pre-culture time,the concentration of agro-infection,infect time,co-culture time and addition of AS) on genetic transformation frequency of Populus 84 K was studied,with the rate of kanamycin resistance seedling as the measure index.The results demonstrated that the optimal differentiation medium and proportion of hormone for leaf-explant was MS BA 0.5 mg/L NAA 0.05 mg/L,and the optimal rooting medium and proportion of hormone for shoot was 1/2 MS NAA 0.05 mg/L IBA 0.05 mg/L.The optimal prioritization optimal system was as follows:to use MS dilute activated agro-infection for 10 times,to infect 5 minutes and to co-culture for 4 days.By adopting above prioritization system of regeneration and genetic transformation,every leaf of populus 84 K could be induced 20 buds on average,rooting rate achieved 99%,rooting number was 6,and transforming rate of leaf disc was 35%.