摘要
目的构建异柠檬酸脱氢酶2(IDH2)野生型/突变型基因慢病毒载体。方法构建IDH2野生型、突变型质粒,采用聚合酶链反应(PCR)进行鉴定;对感染293T细胞,测定病毒滴度;对感染HEK293细胞,观察感染效率。结果PCR鉴定结果显示,2组重组质粒在1.0~2.0kb间各出现一条明显扩增带,片段大小与IDH2目的片段大小一致(1359bp)。对感染293T细胞观察发现,48h后绿色荧光的强度和范围明显增加;病毒滴度为:1.0×10^8TU/mL。感染HEK293细胞的效率在90%以上。结论IDH2野生型和突变型慢病毒构建成功,能够高效地感染细胞。
Objective To construct a lentivirus vector containing IDH 2 wild type/mutant gene.Methods IDH2 wild-type and mutant plasmids were constructed and identified by PCR.293T cells were infected and viral titers were determined.HEK293 cells were infected to observe the infection efficiency.Results The results of PCR showed that there was an obvious amplification band between 1.0-2.0 kb in the two groups,and the size of the fragment was the same as that of the target fragment of IDH 2(1 359 bp).The intensity and range of green fluorescence increased significantly 48 h after infection with 293T cells,and the viral titer was 1.0×10^8 TU/mL.The infection rate of HEK293 cells was more than 90%.Conclusion The wild and mutant lentiviruses of IDH2 have been successfully constructed and can infect cells efficiently.
作者
王世雄
乔健英
刘雪雪
曹相玫
WANG Shixiong;QIAO Jianying;LIU Xuexue;CAO Xiangmei(Grade 2016 in Clinical Medicine Specialty,College of Clinical Medicine;Grade 2016 in Biotechnology Specialty,College of Basic Medicine;Department of Pathology,Basic Medical College Ningxia Medical University,Yinchuan,Ningxia 750004,China)
出处
《现代医药卫生》
2019年第19期2929-2931,2935,共4页
Journal of Modern Medicine & Health
基金
国家自然科学基金项目(81560501)
宁夏回族自治区人力资源和社会保障厅2017留学人员创新创业项目择优资助项目(宁人社函〔2017〕84号)
宁夏回族自治区教育厅大学生创新训练项目(NXCX2018144)
关键词
异柠檬酸脱氢酶
突变型
野生型
肿瘤
慢病毒
包装
Isocitrate dehydrogenase
Mutant type
Wild type
Tumor
Lentivirus
Packaging
