农杆菌介导的天绿香遗传转化影响因素的研究

通过农杆菌介导的方法将新疆准噶尔小胸鳖甲抗冻蛋白基因Mpafp149导入天绿香中,对影响农杆菌转化频率的各种因素进行了研究。结果表明农杆菌介导天绿香遗传转化的潮霉素最佳筛选浓度为8 mg·L^-1，预培养时间2 d，共培养时间2 d，菌液浓度OD₆₀₀为0.2～0.25，浸染时间为8 min时适宜天绿香不定芽的诱导，有利于天绿香转基因植株的获得。通过PCR检测，初步证明外源基因Mpafp149已整合到天绿香基因组中。

The Transformation of Insect Antifreezing Protein for Sauropus androgynus Mediated by Agrobacterium

In order to improve the S. androgynus cold tolerance, genetic transformation system of S. androgynus was established in this study. The cDNA Mpafp149 of Microdera punctipennis dzungarica was used as target gene to obtain the cold tolerant transgenic S. androgynus plants. The stems of S. androgynus were transformed with Agrobacterium tumefaciens EHA105 strain which harbors plasmid pCAMBIA-1302-Mpafp149.The transformation parameters of Agrobacterium concentration, pre-culture period, co-culture period, immersion time and concentrations of antibiotics(hygromycin) were optimized selectively. The results showed that the better physiological and transforming condition was pre-cultured and co-cultured for 2 days respectively, immersed in OD₆₀₀ 0.2~0.25 liquid bacterium for 8 min. It was primarily proved by PCR that the Mpafp149 was integrated into the Sauropus androgynus and the T₀ transgenic S. androgynus plants were obtained successfully.