二倍体草莓匍匐茎遗传转化体系的建立

Establishment of genetic transformation systemusing stolon segments of diploid strawberry (Fragaria vesca)

【目的】为了建立一种可行的草莓稳定遗传转化体系。【方法】以二倍体草莓(Fragaria vesca,‘Hawaii-4’)匍匐茎的茎段为外植体,利用农杆菌介导法进行侵染,并对农杆菌侵染液的浓度、真空渗透时间、暗培养时间和抗生素浓度对草莓茎段转化体系进行了探讨。【结果】以镁离子螯合酶H亚基为操纵基因研究表明,农杆菌侵染液浓度OD600=1,真空渗透时间20 min,暗培养3 d,卡那霉素筛选浓度为35 mg/L为二倍体草莓匍匐茎茎段的最佳遗传转化条件,转化效率达到8%以上。【结论】利用草莓匍匐茎为外植体建立了一种稳定有效的遗传转化转基因体系,为今后草莓的基因改良提供技术支撑。

【Objective】This study is to establish a feasible stable genetic transformation system of strawberry. 【Methods】Stolon stem-segments of diploid strawberry(Fragaria vesca, ’Hawaii-4’) were used as explants to establishthe Agrobacterium-mediated transformation by optimization conditions, including concentrations of Agrobacterium infection solution, vacuum infiltration time, dark culture time, and antibiotic concentration. 【Results】Using magnesium chelatase H subunit(FvCHLH) gene as a reporter gene, the results showed that the optimal parameters for stolon stem-segment transformation were as follows: the concentration of Agrobacterium inoculation solution of OD600 = 1, vacuum infiltration time for 20 min, dark culture under 3 d, and kanamycin screening concentration of 35 mg/L. The transformation efficiency was over 8%. 【Conclusion】Using of strawberry stolon as explants, we have established a stable effective transgenic transformation system, which provides technical support for strawberry genetic improvement in the future.