农杆菌介导的药百合鳞片遗传转化体系的建立

Establishment of an Agrobacterium tumefaciens-mediated genetic transformation system of Lilium speciosum

本试验以药百合鳞片高频再生体系为基础,研究了农杆菌介导的药百合鳞片遗传转化的几个影响因素。结果表明,预培养5d有利于转化;共培养3d并且添加100μmol/L乙酰丁香酮有利于提高转化频率并抑制了农杆菌的过度生长;在侵染阶段添加150μmol/L乙酰丁香酮,农杆菌侵染液pH值5.7、光吸收值0.8,侵染时间10min为最佳遗传转化体系。再生植株经gus基因的瞬时表达检测及hpt基因PCR检测证明载体已成功整合到药百合基因组中。

Several factors,affecting Agrobacterium tumefaciens-mediated genetic transformations of lily were studied based on the establishment of regeneration systems.The pre-culture time of 5 days was beneficial for improving transformation frequency.Co-culture for 3 days with 100 μmol/L AS（acetosyringone） resulted not only in higher rates of transformation but also controlled the over-growth of bacteria.The 10-min bacteria infection（ABS600= 0.8,pH=5.7） with 150 μmol/L AS was found to be the optimal selection for transgenic plantlets.It was confirmed by using gus instantaneous expression test and hpt gene PCR test that the vector was integrated into the regeneration genome of lily.