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根癌农杆菌介导‘粗枝’大叶黄杨P5CS基因转化扶芳藤的研究

Studies on Transformation of Euonymus fortunei with P5 CS Gene Cloned from Euonymus japonicus‘Cu Zhi’ via Agrobacterium tumefaciens

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【作者】 马文慧张倩尚爱芹赵梁军

【Author】 MA Wen-hui~1 ZHANG Qian~(1,2) SHANG Ai-qin~(1,3) ZHAO Liang-jun~1 (1 Department of Ornamental Horticulture and landscape Architecture,China Agricultural University,Beijing 100093; 2 Flower Research Institute,Yunnan Academy of Agriculture Science,Kunming 650205; 3 College of Horticulture,Hebei Agricultural University,Baoding 071001)

【机构】 中国农业大学观赏园艺与园林系云南省农科院花卉研究所河北农业大学园艺学院

【摘要】 采用根癌农杆菌(Agrobacterium tumefaciens,LBA4404菌株)介导法,以‘粗枝’大叶黄杨(Euonymus japonicus‘Cu Zhi’,别名北海道黄杨)的△1-吡咯啉-5-羧酸合成酶基因(P5CS)转化扶芳藤(Euonymus fortunei)的下胚轴。研究了农杆菌菌液浓度、侵染时间、共培养时间、乙酰丁香酮(AS)和PPT浓度(phosphinothricin)等因素对遗传转化的影响。结果表明:根癌农杆菌浓度OD600=0.5,侵染30min,共培养3d,共培养基中添加50μmol/L AS,有利于获得较高的遗传转化效率。在PPT浓度为0.5mg·L-1的分化培养基中筛选到4株抗性植物,经PCR检测初步表明目的基因已经整合到扶芳藤基因组中。

【Abstract】 1- pyrroline-5-carboxylate synthetase(P5 CS) gene cloned from Euonymus japonicus‘Cu Zhi’ was transferred into hypocotyls of Euonymus fortunei mediated by Agrobacterium tumefaciens(LBA4404).Several factors affected genetic transformation such as concentrations of AS and phosphinothricin,the time of infection and co-cultivation were studied.The transformation frequency was improved when the hypocotyls were infected for 30 min with the concentration of Agrobacterium tumefaciens OD600=0.5,and then co-cultivated for 3d on MS medium supplemented with 100μmol/L AS.Four transgenic plants were obtained through phosphinothricin screening on differentiation medium supplemented with 0.5 mg·L-1 PPT.PCR analysis indicated that the P5CS gene integrated into the genome of transgenic plants.

【基金】 国家博士点基金项目(20060019043);北京市科技计划项目(H020620110130)
  • 【会议录名称】 中国观赏园艺研究进展 2009
  • 【会议名称】中国园艺学会观赏园艺专业委员会2009年学术年会
  • 【会议时间】2009-08-18
  • 【会议地点】中国浙江杭州
  • 【分类号】S687
  • 【主办单位】中国园艺学会观赏园艺专业委员会、国家花卉工程技术研究中心
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