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中国生物工程杂志

China Biotechnology
China Biotechnology  2017, Vol. 37 Issue (5): 52-58    DOI: 10.13523/j.cb.20170507
    
Construction of CHO Cell Tably Expressing GLP-1 Analogue and Study of Its Culture Process
ZHANG Jing-jing1, LIU Ke-dong1, QIAN Kai2, MIAO Ya-na1, CAI Yan-fei1, LI Cheng-yuan3, CHEN Yun1, JIN Jian1
1. School of Pharmaceutical Sciences, Jiangnan University, Wuxi 214122, China;
2. Wenzhou University, Wenzhou 325000, China;
3. WuXi Biologics(Shanghai) Co., Ltd, Shanghai 200131, China
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Abstract  

Glucagon like peptide-1 (GLP-1) is a potential drug for the treatment of type II diabetes. However, it has a short half-life in vivo and lot-to-lot expression instability, which limit its clinical application. In our early study, the modified fusion protein NGGH[6×His-tag+Ek+2×GLP-1(A2G)+HSA] was successfully expressed in CHO cells using vector pMH3. The eukaryotic expression plasmid pcDNA3.1/NGGH was constructed and transfected to CHO cells by electroporation. After G418 resistant pressure screening, single cell clone was selected by a new Celigo Imaging Cytometer to obtain stable and high-expression cell clones. The results of Western blot showed that the fusion protein has both GLP-1 and HSA immunogenicity. After batch culture, a stable and high-expression clone was obtained and the yield was 58mg/L. Recombinant cells were cultured in 5L AP20 torrent bioreactor. The result showed that the optimal conditions were pH6.8~7.4 and DO controlled in two-phase way. In these conditions, the yield of NGGH protein rearched 148mg/L.



Key wordsLarge scale culture technology      Glucagon like peptide-1      Chinese hamster ovary cell      Monoclonal screening     
Received: 29 November 2016      Published: 25 May 2017
ZTFLH:  Q819  
Cite this article:

ZHANG Jing-jing, LIU Ke-dong, QIAN Kai, MIAO Ya-na, CAI Yan-fei, LI Cheng-yuan, CHEN Yun, JIN Jian. Construction of CHO Cell Tably Expressing GLP-1 Analogue and Study of Its Culture Process. China Biotechnology, 2017, 37(5): 52-58.

URL:

https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20170507     OR     https://manu60.magtech.com.cn/biotech/Y2017/V37/I5/52

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