Construction of a Differentiated Embryo Chondrocyte 1 Lentiviral Expression Vector and Establishment of its Stably Trans
Human differentiated embryo chondrocyte 1 (DEC1), has been suggested to play key roles in immune regulation, cell differentiation, proliferation and apoptosis, circadian rhythms, hypoxia response and carcinogenesis. However, the role of DEC1 in gastric ca
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Construction of a Differentiated Embryo Chondrocyte 1 Lentiviral Expression Vector and Establishment of its Stably Transfected HGC27 Cell Line Rui Hu, Yun-Shan Wang, Yi Kong, Pin Li, Yan Zheng, Xiao-Li Ma and Yan-Fei Jia Abstract Human differentiated embryo chondrocyte 1 (DEC1), has been suggested to play key roles in immune regulation, cell differentiation, proliferation and apoptosis, circadian rhythms, hypoxia response and carcinogenesis. However, the role of DEC1 in gastric cancer have not been well established. Lentiviral vectors are widely used for the stable expression of genes and currently under development for clinical use in gene therapy. Therefore we intended to construct a lentiviral DEC1 expression vector and then establish a gastric cancer cell line with stable expression of DEC1. The coding sequence of gene was amplified using PCR and cloned into pGV218 vector. 293T cells were transfected using Lipofectamine 2000 and packaged for the recombinant lentivirus particles. When the cloned sequence was identified to be right, the recombinant lentivirus particles were amplified in a large quantity. The titer of virus was determined by real-time PCR. Subsequently, we collected the lentivirus venom to infect the HGC27 cells and establish a stable, overexpressed cell line named GFP/DEC1-HGC27. This study will provide a new cell model for further study of the role of DEC 1in the pathogenesis of gastric cancer. Keywords DEC1
Lentiviral expression vector Gastric cancer HGC27
R. Hu Y.-S. Wang Y. Kong P. Li Shandong University School of Medicine, Jinan, Shandong 250012, China Y.-S. Wang Y. Zheng X.-L. Ma Y.-F. Jia (&) Central Laboratory, Jinan Central Hospital, Affiliated to Shandong University, Jie fang road 105, Shandong, Jinan, China e-mail: [email protected]
S. Li et al. (eds.), Frontier and Future Development of Information Technology 2599 in Medicine and Education, Lecture Notes in Electrical Engineering 269, DOI: 10.1007/978-94-007-7618-0_325, Springer Science+Business Media Dordrecht 2014
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325.1 Introduction Human differentiated embryo chondrocyte 1 (DEC1), a basic helix-loop-helix (bHLH) transcription factor, has rat and mouse orthologs named enhancer of split and hairy related protein-2 (SHARP-2) and stimulation of retinoic acid 13 (Stra13), respectively [1]. Overexpression of DEC1 has been reported to contribute to cell differentiation, proliferation and apoptosis, circadian rhythms, hypoxia response and carcinogenesis [1–4]. High levels of DEC1 expression are also detected in a variety of cancer cell lines including leukaemia, colon and lung adenocarcinoma, gliomas and breast cancer [5–7] indicating it play a pivotal role in carcinogenesis. We have previously shown that DEC1 mRNA and protein were upregulated significantly in gastric cancer tissue [4]. Importantly, the increased DEC1 expression was correlated with poor differentiation of gastric cancer, suggesting that DEC1 may play an important role in the differentiation and progression of gastric tumors. Ho
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