摘要
【目的】构建转血管内皮生长因子165(VEGF165)基因的红花,为利用植物生物反应器规模化生产VEGF165奠定基础。【方法】采用PCR方法,克隆获得植物偏好性的VEGF165基因,将其与pOP质粒连接,构建重组质粒pOP-VEGF165,对其进行NcoⅠ和HindⅢ双酶切及PCR鉴定。采用冻融法将质粒pOP-VEGF165转入根癌农杆菌EHA105中,通过农杆菌介导获得转化红花植株,经PCR鉴定得到转基因红花阳性植株。【结果】克隆获得了495bp的VEGF165基因片段,并成功构建了植物特异表达载体pOP-VEGF165,用其转化红花子叶后,共获得了8株转基因红花植株,其中2株鉴定为阳性。【结论】获得了转VEGF165基因的红花株系。
【Objective】This study constructed transgenic safflower with human vascular endothelial growth factor(VEGF165)gene to improve the large-scale production of VEGF165 using plant bioreactor.【Method】Plant-preferred VEGF165 gene was cloned by PCR and connected with pOP vector to form recombinant vector pOP-VEGF165.The recombinant vector was then identified by double digestion with NcoⅠ and Hind Ⅲ.Freeze-thaw method was used to transfer plasmid pOP-VEGF165 into Agrobacterium tumefaciens EHA105 and safflower cotyledons were obtained by Agrobacterium mediation.The positive transgenic plants were determined by PCR analysis at last.【Result】VEGF165gene fragment with length of 496 bp was cloned and it was used to construct to plant specific expression vector pOP-VEGF165.A total of 8positive transgenic safflower plants were obtained.【Conclusion】Safflower lines with VEGF165 gene were obtained successfully.
出处
《西北农林科技大学学报（自然科学版）》
CSCD
北大核心
2014年第11期146-150,共5页
Journal of Northwest A&F University(Natural Science Edition)
基金
2013年国家大学生创新创业训练计划项目(201310193050)
国家高新技术研究与发展计划项目(2011AA100606)
教育部博士点基金青年教师基金项目(20122223120002)
吉林省中医药管理局项目(2012162)
吉林省教育厅项目(201458)
吉林农业大学2012年校内博士启动基金项目(201207)
