农杆菌介导的大豆子叶节和下胚轴转化方法的比较及优化

Comparison and Optimization of the Agrobacterium-mediated Transformation of Soybean by Using Cotyledonary Node and Hypocotyl Explants

以东农50、东农42和黑农44的子叶节和下胚轴为外植体,对农杆菌介导的大豆子叶节和下胚轴2种转化方法进行了比较和优化。结果表明:不同转化方法对大豆的品种要求不同,东农50适合子叶节转化法,而黑农44适合下胚轴转化法。在子叶节转化体系中,卡那霉素的筛选浓度是100 mg.L-1,在下胚轴转化体系中,卡那霉素的筛选浓度是75 mg.L-1,可见下胚轴体系比子叶节体系在卡那霉素筛选过程中表现得更为敏感。在子叶节和下胚轴转化体系中,最适乙酰丁香酮浓度均为200 mol.L-1,最适共培养时间均为3 d。在子叶节转化体系中,5 d苗龄时转化率最高,而下胚轴转化体系中,4～6 d苗龄时转化率均较高,以6 d苗龄最高。

Soybean cotyledonary node and hypocotyl transformation methods were compared and optimized using three soybean genotypes,Dongnong 50,Dongnong 42 and Heinong 44.The results showed that different transformation methods required different soybean varieties.Dongnong 50 was suitable to cotyledonary node transformation method,while Heinong 44 was suitable to hypocotyl transformation method.In the cotyledonary node transformation system,the screening concentration of Kanamycin was 100 mg·L-1,while in the hypocotyl transformation system was 75 mg·L-1,the hypocotyl transformation system was more sensitive than the cotyledonary node transformation system to kanamycin.The optimal concentration of acetosyringone was both 200 mol·L-1,the optimal co-culture time was both 3 d in the cotyledonary node and hypocotyl transformation systems.In the cotyledonary node transformation system,conversion rate was the highest when the seedling age was 5 d,while in the hypocotyl transformation system,conversion rate was higher in 4-6 d,in which the highest was 6 d.