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《China Biotechnology》 2015-05
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Construction of A Cell Line EGFP-Luc-Hepa1-6 and Its Application in the Mouse Model of the Hepatoma

YE Yu-chen;ZHAO Jun-long;WANG Lin;DUAN Juan-li;GAO Chun-chen;QIN Hong-yan;DOU Ke-feng;Department of Hepatobiliary Surgery,Xijing Hospital;State Key Laboratory of Cancer Biology,Department of Medical Genetics and Developmental Biology,Fourth Military Medical University;  
Objectives: We aimed to construct an eukaryotic expressive vector p CMV-Luciferase-IRES2-EGFP which contains luciferase and enhanced green fluorescent protein( EGFP) and steadily screen Hepa1-6 cell line transfected with this recombinant vector,with the purpose of stably labeling and in vivo tracking the hepatoma cells in orthotopic hepatoma model established using this cell line. Methods: p GL3-Basic vector was digested by Xba I,end-repaired by Klenow Fragment and then redigested by Xho I to acquire the luciferase gene,which was ligated with the larger fragment of p IRES2-EGFP vector through Xho I / Sma I double digestion to construct the recombinant vector. Authenticated by endonuclease digestion and sequencing,the recombinant product presented the correct sequence. The expression of EGFP and activity of luciferase of Hepa1-6 cells transfected with this vector were,respectively,dectected by fluorescent microscope and dual-luciferase reporter assay and Lumina Ⅱimaging system. Confirming the expression of the vector in the cell line,we inoculated this cell line orthotopically in the liver of C57 BL /6 mouse to establish a hepatoma model. The growth of the hepatoma was consecutively monitored in vivo using Lumina Ⅱ imaging system and both paraffin sections( HE staining) and frozen sections(immunofluorescent staining) were prepared from the ex vivo tumor tissue to observe the pathological features and EGFP expression of the hepatoma cells. Results: Hepa1-6 cell line expressing the recombinant vector p CMVLuciferase-IRES2-EGFP( EGFP-Luc-Hepa1-6) was successfully constructed. Using this cell line,an orthotopic hepatoma model in mice was established with the capability of in vivo tracking and ex vivo labeling the hepatoma cells. Conclusions: The EGFP-Luc-Hepa1-6 cell line was constructed successfully and the orthotopic hepatoma model established using this cell line can satisfy the needs of detecting the progression of the hepatoma both in vivo and ex vivo.
【Fund】: 国家自然科学基金重大研究计划培育项目(91029731) 国家自然科学基金青年科学基金(81100307) 国家自然科学基金青年科学基金(81401940);; 肿瘤生物学国家重点实验室(第四军医大学)开放基金(Cbskl201408)资助项目
【CateGory Index】: R735.7;R-332
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【Citations】
Chinese Journal Full-text Database 1 Hits
1 YANG Qing, YANG Guangshun, WEI Lixin, JIA Fengqi, WANG Weifeng, WU Mengchao, GUO Yajun. Tumor Immunology and Biotherapy Center, Eastern Institute of Hepatobiliary Surgery, Second Military Medical University, Shanghai 200438, China;The in vivo antitumor activity of murine liver tumor vaccine expressing MIP-1α[J];Chiese Journal of Surgery;2002-10
【Co-citations】
Chinese Journal Full-text Database 10 Hits
1 Sasipawan Machana;Natthida Weerapreeyakul;Sahapat Barusrux;Kanjana Thumanu;Warapom Tanthanuch;Graduate School,Faculty of Pharmaceutical Sciences,Khon Kaen University;Center for Research and Development of Herbal Health Products,Faculty of Pharmaceutical Sciences,Khon Kaen University;Centre for Research and Development of Medical Diagnostic Laboratories,Faculty of Associate Medical Sciences,Khon Kaen University;Synchrotron Light Research Institute(Public Organization),Nakhon Ratchasima;;Synergistic anticancer effect of the extracts from Polyalthia evecta caused apoptosis in human hepatoma(HepG2)cells[J];亚太热带生物医学杂志(英文版);2012-08
2 Sasipawan Machana;Natthida Weerapreeyakul;Sahapat Barusrux;Graduate School,Faculty of Pharmaceutical Sciences,Khon Kaen University;Center for Research and Development of Herbal Health Product,Faculty of Pharmaceutical Sciences,Khon Kaen University;Center for Research and Development of Medical Diagnostic Laboratories,Faculty of Associate Medical Sciences,Khon Kaen University;;Anticancer effect of the extracts from Polyalthia evecta against human hepatoma cell line(HepG2)[J];亚太热带生物医学杂志(英文版);2012-05
3 YI Yong-xiang;ZHAO Wei;HAN Jian-bo;Second Affiliated Hospital of Southeast University;;Observation of the effect of tumor model establishment for hepatocellular carcinoma in mice via tumor cell intrahepotic injection[J];Journal of Hepatopancreatobiliary Surgery;2013-05
4 FENG Yingqiao;YANG Yuansheng;CUI Shulan;CHEN Ken;WANG Hui;School of Clinical Medicine,Guangdong Pharmaceutical University;The Second Affiliated Hospital,Guangdong Pharmaceutical University;School of Nursing,Guangdong Pharmaceutical University;School of Traditional Chinese Medicine,Guangdong Pharmaceutical University;;Advance in the development and application of experimental animal models of liver fibrosis[J];Journal of Guangdong Pharmaceutical University;2013-05
5 ZHOU Chenjie;GONG Xumeng;CAI Liquan;WANG Yan;GAO Yi;Second Department of Hepatobiliary Surgery,Zhujiang Hospital;;A modified approach to establishing a rat model of liver cancer with concurrent cirrhosis[J];Journal of Southern Medical University;2014-04
6 WANG Yi-Tian;ZHANG Meng;WANG Bo;CAI Zhi-Ming;WU Song;Clinical Medical College, Shenzhen Second People's Hospital, Anhui Medical University;Zhongshan School of Medicine, Sun Yat-sen University;;Research progress of telomerase reverse transcriptase in urogenital carcinoma[J];Chinese Bulletin of Life Sciences;2014-08
7 ZHU Xin-hong;MA Gui-liang;MAO Wei-zheng;LI Qian;ZHAN Shu-hui;ZHOU Shao-fei;LIN Cun-zhi;Qingdao Municipal Hospital;Department of Biology Treatment for Respiratory Oncology,Affiliated Hospital of Qingdao University;;Experimental study on mice gastric cancers treated by oncolytic virus with transgenic TNF-αexpressed in umbilical cord blood MSCs[J];Chinese Journal of Cancer Prevention and Treatment;2015-05
8 ZHOU Min,YANG Dong-hua. Department of Gastroenterology,First Affiliated Hospital,Jinan University,Guangzhou 510630,China;Current status and progress of tumor vaccine for hepatocellular carcinoma.[J];Chinese Journal of Hepatology;2003-12
9 Jia Yunlong;Wang Yu;Wang Tingting;Duan Yuqing;Ma Ming;Wang Miao;Wang Hongyan;Liu Lihua;Department of Biotherapy,the Fourth Hospital of Hebei Medical University;Department of Chest Surgery,the Fourth Hospital of Hebei Medical University;;Analysis on lymphocyte subsets in tumor-draning lymph nodes of esophageal cancer patients[J];Chinese Journal of Cancer Biotherapy;2014-02
10 WANG Tingting;WANG Yu;JIA Yunlong;WANG Xuexiao;WANG Jiali;LV Wei;DUAN Yuqing;LIU Lihua;Department of Biotherapy, The Fourth Hospital of Hebei Medical University;;Effects of Adoptive Immunotherapy on T Cell Subsets of Peripheral Blood in Non-small Cell Lung Cancer Patients[J];Cancer Research on Prevention and Treatment;2015-04
【Secondary Citations】
Chinese Journal Full-text Database 1 Hits
1 JIA Feng Qi, WANG Wei Feng, JIANG Xiao Ling, YANG Qing, WEI Li Xin, YANG Guang Shun, WU Meng Chao (Tumor Immunology and Biotherapy Center, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438, China);Construction of adenoviral vector carrying murine MIP1-α and its expression in hepatocellular carcinoma[J];Academic Journal of Second Military Medical University;2002-05
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