以PMI为选择标记基因的雪柑遗传转化体系的优化  

Optimization of Genetic Transformation System of 'Xuegan' Based on the PMI Selection System

作　　者：王会全[1] 吴少华[2] 余志雄[1] WANG Hui-quan;WU Shao-hua;YU Zhi-xiong(Fujian Vocational College of Agriculture,Fuzhou,Fujian 350119;Institute of Natural Products of Horticultural Plants,Fujian Agriculture and Forestry University,Fuzhou,Fujian 350002)

机构地区：[1]福建农业职业技术学院,福建福州350119 [2]福建农林大学园艺植物天然产物研究所,福建福州350002

出　　处：《安徽农业科学》2018年第36期83-89,共7页Journal of Anhui Agricultural Sciences

基　　金：福建省教育厅青年基金项目(JA12423);福建农业职业技术学院青年项目(2016JS0004)

摘　　要：[目的]以PMI为选择标记基因对雪柑的遗传转化体系进行优化。[方法]对农杆菌的菌液浓度、实生苗培养条件、甘露糖筛选压、2,4-D浓度、2,4-D预处理时间、农杆菌侵染时间、共培养时间进行梯度试验来研究转化后的再生率,并对筛选培养基中6-BA和NAA不同浓度组合进行试验来研究抗性大苗的筛选。[结果]经过暗培养20 d光培养10 d条件的实生苗外植体在农杆菌菌液OD600为0.6,1.0 mg/L 2,4-D预处理3 h,农杆菌侵染30 min,共培养时间4 d,筛选压为甘露糖20 g/L的情况下转化后的再生率大大提高,而在筛选培养基中加入2 mg/L 6-BA+0.5 mg/L NAA有助于抗性大苗的筛选。[结论]通过对影响转化的各种因素的优化试验建立了雪柑以PMI基因/甘露糖选择标记系统的高效遗传转化体系。[Objective] To optimize the genetic transformation system of Citrus using PMI as a selective marker gene.[Method] Gradient experiments were carried out to study the regeneration frequency of ‘Xuegan’by the concentration of bacterial solution,culture condition of seedlings,screening pressure of mannose,concentration of 2,4-D,pretreatment time of 2,4-D,infection time of Agrobacterium and co-culture time. The selection of resistant plantlets with different concentrations of 6-BA and NAA in screening medium was studied. [Result] The seedling explants cultured under dark culture for 20 days and light culture for 10 days were pretreated with Agrobacterium solution OD600 for 0.6,1.0 mg/L,2,4-D for 3 hours,infected with Agrobacterium for 30 minutes,co-cultured for 4 days,and pressed with mannose for 20 g/L. The regeneration frequency of transformed seedlings was greatly increased,while 2 mg/L 6-BA+0.5 mg/L NAA added to the screening medium was helpful for screening resistant seedlings. [Conclusion] An efficient genetic transformation system of‘Xuegan’with PMI gene/mannose selective marker system was established by optimizing the factors affecting transformation.

关 键 词：柑橘 6-磷酸甘露糖异构酶 遗传转化 

分 类 号：S188[农业科学—农业基础科学]