| 引用本文: | 吴洁,李剑波,高孟,金素凤,潘海桦,陈刚,姜云水,庄昉成.HEK293细胞的传代、建库与高效表达重组HPV16-E6E7腺病毒的研究[J].中国现代应用药学,2015,32(4):399-403. |
| WU Jie,LI Jianbo,GAO Meng,JIN Sufeng,PAN Haihua,CHEN Gang,JIANG Yunshui,ZHUANG Fangcheng.Study on HEK293 Cell to Passage, Establish Cell Banks, and Highly Efficiency Express a Recombinant HPV16-E6E7 Adenovirus Vector[J].Chin J Mod Appl Pharm(中国现代应用药学),2015,32(4):399-403. |
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| 摘要: |
| 目的 对HEK293细胞进行三级细胞库的建立，并高效表达重组的HPV16-E6E7腺病毒。方法 进行HEK293细胞传代培养，并建立三级细胞库，按中国药典2010年版III部要求进行常规检验；培养重组HPV16-E6E7腺病毒，测定滴度，插入目的基因以及基因组酶切图谱鉴定。结果 HEK293细胞传代符合其生长形态，建立的三级细胞库和不同细胞库之间的传代数，细胞浓度和装量均符合疫苗基质制备的要求；能高效表达重组HPV16-E6E7腺病毒，种子滴度>3.0×109 IU?mL-1；表达的目的基因和目的蛋白稳定。结论 建立的HEK293三级细胞库符合疫苗用细胞基质的要求。 |
| 关键词: HEK293 细胞库 HPV16 重组腺病毒 高效表达 |
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| 基金项目:浙江省科技计划项目(2008F3022)；杭州市科技局重大科技创新项目(20112313A37) |
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| Study on HEK293 Cell to Passage, Establish Cell Banks, and Highly Efficiency Express a Recombinant HPV16-E6E7 Adenovirus Vector |
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WU Jie1, LI Jianbo1, GAO Meng1, JIN Sufeng2, PAN Haihua2, CHEN Gang1, JIANG Yunshui1, ZHUANG Fangcheng1
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1.Institute of Viral Diseases,Hangzhou 310013, China;2.Zhejiang Pukang Biotechnology Co., Ltd., Hangzhou 310053, China
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| Abstract: |
| OBJECTIVE To study on HEK293 cell to passage, establish 3-level cell banks, and by that to highly efficiency express a recombinant HPV16-E6E7 adenovirus vector. METHODS HEK293 cell passaged and established 3-level cell banks. And general inspected according to the requirements of Chinese Pharmacopeia Ⅲ 2010. By the HEK293 to propagate a recombinant HPV16-E6E7 adenovirus vector, identitied its titer, inserted target gene and genome macrorestriction. RESULTS HEK293 cell met the growthform of its description; the limited passages, concentration of cells, and install quantity of 3-level cell banks were all conformed to requirements of vaccine substrates; the expressed recombinant HPV16-E6E7 adenovirus titer was more than 3.0×109 IU?mL-1; the expressed inserted target gene and genome macrorestriction map were stable. CONCLUSION HEK293 cell and its 3-level cell banks established are all conformed to the requirements of vaccine substrates. |
| Key words: HEK293 cell bank HPV16 recombinant adenovirus highly efficiency expression |
