农杆菌介导韭菜遗传转化相关因素的研究

Research on the Factors Related to Agrobacterium-mediated Genetic Transformation of the Chinese Chive

以"汉中冬韭"韭菜品种为试验材料,用含有pCAMBIA3301质粒的根癌农杆菌菌株EHA105对影响韭菜遗传转化效率的多种因素进行研究。结果表明,诱导40 d的愈伤组织,GUS基因瞬时表达率达到93%,且最适宜于不定芽的分化;当乙酰丁香酮(AS)的浓度为100μmol/L时,愈伤的GUS表达率达到91%,植株再生率为7.9%,AS浓度增加时其值也不会增加;菌液OD600值为0.6侵染10 min时,与其他组合相比,外植体受伤程度小,GUS表达率及再生率最高;侵染后的愈伤共培养3 d后,农杆菌生长较少,GUS表达率为91.1%,而再生率达到7.2%,为最佳的共培养时间。通过试验得到韭菜遗传转化因素的最佳条件,为今后的遗传转化提供一些参考。

Using ＂Hanzhong Dongjiu＂ variety and the Agrobacterium EHA105 containing pCAMBIA3301 plasmid as the material, it studied the factors which could influence transformation efficiency of Chinese chive. The results showed that transient expression rate of GUS gene reached 93% with the callus cultured for 40 d as the explant and the adventitious buds differentiated well. When 100 μmol/L acetosyringone was added to the miedium, GUS expression rate of the callus reached 91% and the plant regeneration rate was 7.9%, but did not increase with increasing of the AS concentration. Compared with other combinations, the calli were soaked in Agrobacterium （ OD600 - 0.6 ） for 10 min, the injury degree of explants was slight and the GUS expression rate and regeneration rate were the highest. When the calli were co-cuhured for 3 days after infection, the bacterium grew less than other period, GUS expression rate and the regeneration rate were 91.1% and 7.2% respectively. Through the experiment, the best conditions of genetic transformation factors for chinese chive were obtained,