Please wait a minute...

中国生物工程杂志

CHINA BIOTECHNOLOGY
中国生物工程杂志  2011, Vol. 31 Issue (11): 75-80    
技术与方法     
稳定表达人Siat7e基因的MDCK细胞系的建立和全悬浮细胞的驯化
张妍, 张文俊, 李群辉, 刘晓文, 刘文博, 刘秀梵
扬州大学兽医学院 农业部畜禽传染病学重点开放实验室 扬州 225009
Generation of a MDCK Cell Line to Suspension Culture by Stably Experessing Human Siat7e Gene
ZHANG Yan, ZHANG Wen-jun, LI Qun-hui, LIU Xiao-wen, LIU Wen-bo, LIU Xiu-fan
Key Laboratory of Animal Infectious Diseases, Ministry of Agriculture, Yangzhou University, Yangzhou 225009, China
 全文: PDF(595 KB)   HTML
摘要:

目的:现有的禽流感疫苗生产的方法已经不能适应工业化大生产的需求,有必要开发全悬浮培养的细胞系来满足大生产的需求。方法:我们通过转染稳定表达 Siat7e 基因的真核表达载体对MDCK细胞进行改造,并经过后期的驯化,筛选适应于全悬浮培养的MDCK细胞。结果:成功筛选到能稳定表达 Siat7e 基因并能适应全悬浮生长的细胞系。结论:该细胞系具有潜在的应用价值,为MDCK细胞的培养以及工业化大生产疫苗提供参考。

关键词: MDCKSiat7e全悬浮疫苗生产    
Abstract:

Object:Current influenza vaccine manufacturing have been unable to meet the demand of Industrialized production. Method: Anchorage-dependent MDCK cells were converted to anchorage-independent cells, capable of growing in suspension as a result of transfection with the human Siat7e gene (ST6GalNac V). Result:Successfully conversioned a MDCK cell line stably experessing human Siat7e and they were capable of growing in suspension cultures. Conclusion:These results are critical for establishing large-scale suspension culture of MDCK cells as subsequent well as large-scale vaccine production.

Key words: MDCK    Siat7e    Suspension culture    Vaccine manufacturing
收稿日期: 2011-05-20 出版日期: 2011-11-25
ZTFLH:  Q786  
基金资助:

现代农业产业技术体系建设专项资金(nycytx-41-G07)、国家"973"计划(2011CB505003))资助项目

通讯作者: 刘秀梵     E-mail: xfliu@yzu.edu.cn
服务  
把本文推荐给朋友
加入引用管理器
E-mail Alert
RSS
作者相关文章  

引用本文:

张妍, 张文俊, 李群辉, 刘晓文, 刘文博, 刘秀梵. 稳定表达人Siat7e基因的MDCK细胞系的建立和全悬浮细胞的驯化[J]. 中国生物工程杂志, 2011, 31(11): 75-80.

ZHANG Yan, ZHANG Wen-jun, LI Qun-hui, LIU Xiao-wen, LIU Wen-bo, LIU Xiu-fan. Generation of a MDCK Cell Line to Suspension Culture by Stably Experessing Human Siat7e Gene. China Biotechnology, 2011, 31(11): 75-80.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/        https://manu60.magtech.com.cn/biotech/CN/Y2011/V31/I11/75


[1] Tree J A, Richardson C, Fooks A R. Comparison of large-scale mammalian cell culture systems with egg culture for the production of influenza virus A vaccine strains. Vaccine, 2001, 19:3444-3450.

[2] Stohr K, Esveld. Will vaccines be available for the next influenza pandemic? Science, 2004, 306:2195-2196.

[3] Genzel Y, Reichl U. Continuous cell lines as a production system for influenza vaccines. Vaccine, 2009, 8(12): 1681-1692.

[4] Frisch S M, Francis H. Disruption of epithelial cell-matrix interactions induces apoptosis. Cell Biol, 1994, 124:619-626.

[5] Folkman J, Moscona A. Role of cell shape in growth control.Nature,1978,273:345-349.

[6] Tichopad A, Dzidic A, Pfaffl M.Improving quantitative real-time RT-PCR reproducibility by boosting primer-linked amplification efficiency. Biotechnol, 2002, 24:2053-2056.

[7] Akiko T, Tetsuya O, Tsuchida A, et al. Synthesis of disialyl Lewis a structure in colon cancer cell lines by a sialyltransferase, ST6GalNAc VI, responsible for the synthesis of alpha-series gangliosides. Biol Chem, 2003,278:22787-22794.

[8] Jaluria P, Betenbaugh M, Konstantopoulos K, et al. Application of microarrays to identify and characterize genes involved in attachment dependence in HeLa cells. Metab Eng, 2007, 9:241-251.

[9] Hakomori S I. Cell adhesion/recognition and signal transduction through glyco-sphingolipid microdomain. Glycoconj, 2000,17:143-151.

[10] Wittwer C, Herrmann A, Moss A, et al. Continuous fluorescence monitoring of rapid cycle DNA amplification.BioTechniques,1997,22:130-138.

[11] Wiesner R, Ruegg J, Morano I. Counting target molecules by exponential polymerase chain reaction copy number of mitochondrial DNA in rat tissues.Biochem Biophys Res,2002,183:553-559.

[12] Schlereth W, Bassukas D, Deubel W, et al. Use of the recursion formula of the Gompertz function for the quantitation of PCR-amplified templates. Mol Med,1997,1:463-467.

[13] Liu W, Saint D.Validation of a quantitative method for real time PCR kinetics.Biochem,2002,294:347-353.

[1] 刘赛宝,李亚芳,王辉,王伟,冉多良,陈洪岩,孟庆文. 利用CRISPR/Cas9技术构建流感病毒高产细胞系MDCK-Tpl2 -/-*[J]. 中国生物工程杂志, 2019, 39(1): 46-54.
[2] 龚迪, 易小萍, 张元兴. MDCK细胞微载体悬浮培养放大工艺研究[J]. 中国生物工程杂志, 2012, 32(09): 55-60.
[3] 石贵贤. 疫苗生产商开始用连续细胞培养代替滚瓶进行生产[J]. 中国生物工程杂志, 1987, 7(5): 82-82.
[4] 石贵贤. 克隆迫近疫苗生产的AIDS基因[J]. 中国生物工程杂志, 1985, 5(3): 77-77.