超声波直接转导外源基因转化八棱海棠的技术研究  

Study on method of ultrasonic-directed transformation of Malus robusta Rehd

作　　者：丛郁[1] 渠慎春[1] 乔玉山[1] 姚泉洪[2] 章镇[1] 

机构地区：[1]南京农业大学园艺学院,江苏南京210095 [2]上海市农业遗传重点实验室/上海市农业科学院生物技术研究中心,上海201106

出　　处：《南京农业大学学报》2007年第3期42-46,共5页Journal of Nanjing Agricultural University

基　　金：国家自然科学基金资助项目(30370987);江苏省科技研究资助项目(BJ2003309)

摘　　要：应用超声波直接转导法,对八棱海棠进行rolC基因转化,同时,利用gus基因瞬间表达的方法研究了超声波处理时间、处理功率和转化缓冲液中二甲基亚砜(DMSO)对rolC基因转化率的影响。结果表明:当DMSO为5%(体积分数)时,用80 W功率处理20 min,能够获得最佳的转化效果。在超声波最佳处理条件下转化486个八棱海棠叶片,共得到237个抗性愈伤组织和9株抗性苗,转化率为1.85%。GUS染色、PCR及Southern blotting检测结果显示,有8个八棱海棠株系的基因组中整合了完整的外源rolC基因。Malus robusta Rehd. was transformated with rolC gene by ultrasonic-directed transformation method. The factors which influenced the transformation efficiency, such as sonication treatment time, power and the DMSO in transformation buffer, were all investigated by GUS staining instant expression. The results demonstrated that the optimum treatment was 80 W, 20 minutes and 5% DMSO. 486 leaves were transformated under the optimum treatment condition, then 237 resistant callus and 9 resistant buds were obtained （transformation rate 1.85% ）. Among the 9 resistant buds, 8 resistant buds were verified to be transformants by GUS staining, PCR amplification and Southern blotting, which was suggested that the rolC gene had been integrated into genomic of Malus robusta Rehd..

关 键 词：八棱海棠 超声波转化 ROLC基因 

分 类 号：S603.6[农业科学—园艺学]