【作者】 付洪冰； 张俊华； 崔崇士；

【Author】 FU Hong-Bing,CUI Chong-Shi ZHAO Xi,LIU Qi, (Horticulture College,Northeast Agricultural University,Harbin Helongjiang 150030)

【机构】 东北农业大学园艺学院；

【摘要】 本试验以"金辉一号"南瓜为试材,利用根癌农杆菌介导转化南瓜子叶节,研究了预培养时间、侵染时间、乙酰丁香酮(AS)浓度、共培养时间等因素,以及抗生素羧苄青霉素(Carb)、头孢霉素(Cef)和筛选剂卡那霉素(Kan)对离体不定芽的影响,建立最佳遗传转化体系。结果表明:外植体预培养0天,侵染时间30min,AS浓度100mg/L,共培养时间5天时获得最高遗传转化效率;最适除菌剂为Cef,最适浓度为500 mg/L,最适Kan筛选浓度为100 mg/L;在MS培养基上培养抗性芽生根,经PCR、Southern-blot检测,证明为转基因植株。更多还原

【Abstract】 To establish transformation system is a fundamental role to the transgenic study.Efficient transformation and screening system of Jinhuiyihao(C.mosehata Dueh) mediated by Ag robateriurn was developed.The leaves from tissue culture served as excellent explants for transformation,and many factors affected the transformation frequency,such as pre-culture time of explants,infection time and concentration of acetosyringone(AS) for constructing genetic transformation system.Additionally, co-infection time and the concentration of antibiotics which inhibited the growth of Agrobacterium were also discussed.The data was analyzed by software DPS,and a suitable genetic transformation system was established.The results indicated that pre-culturation for 0 day,infection for 30 minutes,AS with a concentration of 100mg/L and co-infection for 5 days would lead to an obtaining of transformation efficiency.While the medium with Cefotaxime 500 mg/L,kanamycin 100 mg/L was found optimal for selection of transgenic plants.Eventually,the result of Southern blot showed that the kanamycin-resistant exogenous gene was integrated into the genome of cucurbita.更多还原