以对潮霉素抗性为筛选标记的棉花遗传转化

Hygromycin Resistance as an Efficient Selectable Marker for Cotton Stable Transformation

以 HPT基因作为筛选标记基因 ,潮霉素作为筛选剂 ,通过农杆菌介导法将 GFP基因导入棉花细胞并得到再生植株。经过 Southern杂交证实外源基因已经整合到棉花基因中 ,荧光显微镜可以观察到绿色荧光 ,说明 GFP基因得到表达。研究确定了潮霉素作为棉花转化的筛选剂在农杆菌介导的转化中适用的浓度范围及筛选方法 ,即棉花转化中潮霉素的筛选浓度范围为 2 .5～ 1 0 mg· L- 1,在有 GFP等一些易于观察和检测的标记基因存在时 ,起始浓度可用较低 ( 2 .5 mg· L- 1)的筛选浓度 ;而在没有标记基因或检测方法较复杂时 ,起始浓度用较高 ( 1 0 mg· L- 1)

Employing HPT as a selection marker gene and hygromycin as a selection agent, The GFP gene was transformed into cotton via agrobacterium and regeneration plants were acquired. Transgenic plants were confirmed by southern hybridization, expression of the GFP gene was showed with fluorescence microscope. Using hygromycin as a selection agent in the cotton transformation, the reasonable selection concentration of should be 2.5~10 mg·L 1 if marker gene, such as GFP gene , exists or easy to be examined , the start concentration of hygromycin in the medium should be 2.5 mg·L 1 ; If no marker gene exists or the examination is very complicated, the start concentration of hygromycin could be higher as 10 mg·L 1 . So that the higher rate of transformation can be obtained.