| 重组人白细胞介素1受体拮抗蛋白荧光质粒在软骨细胞的表达 |
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| 引用本文： | 张 平,刘 斌,蔡道章,钟志宏,潘永谦,张振山. 重组人白细胞介素1受体拮抗蛋白荧光质粒在软骨细胞的表达[J]. 中国组织工程研究, 2011, 15(7): 1169-1173. DOI: 10.3969/j.issn.1673-8225.2011.07.007 |
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| 作者姓名： | 张 平 刘 斌 蔡道章 钟志宏 潘永谦 张振山 |
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| 作者单位： | 1广州医学院第三附属医院骨外科，广东省广州市5101502中山大学附属第三医院骨外科，广东省广州市 510630 |
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| 基金项目： | 广东省科技计划项目(粤科规划字(2009)198号)，广州医学院博士启动项目(2008C20)。 |
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| 摘 要： | 背景：白细胞介素1受体拮抗蛋白能延缓骨性关节炎进程，通过转基因方法可以使白细胞介素1受体拮抗蛋白表达的增加。目的：观察重组人重组人白细胞介素1受体拮抗蛋白荧光质粒的构建及经脂质体转染软骨细胞的表达情况。方法：双酶切法切取重组人白细胞介素1受体拮抗蛋白的c-DNA片段，通过T4DNA连接酶连接到pEGFP-C1载体上。体外分离培养兔关节软骨细胞，然后用构建的重组人白细胞介素1受体拮抗蛋白质粒经脂质体转染软骨细胞，通过荧光显微镜观察转基因的表达和荧光定量PCR检测其表达。结果与结论：获得重组人pEGFP-C1-IL-1Ra真核表达载体质粒，酶切及测序结果证明表达质粒的DNA序列完全正确。荧光显微镜观察有绿色荧光蛋白表达，荧光定量PCR鉴定证实转染的软骨细胞基因得到表达。
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| 关 键 词： | 白细胞介素1受体拮抗蛋白 基因转染 荧光 质粒 软骨细胞 表达 |
| 收稿时间： | 2010-09-20 |
Expression of recombinant human interleukin-1 receptor antagonist protein fluorescent plasmid in chondrocytes |
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| Zhang Ping,Liu Bin,Cai Dao-zhang,Zhong Zhi-hong,Pan Yong-qian,Zhang Zhen-shan. Expression of recombinant human interleukin-1 receptor antagonist protein fluorescent plasmid in chondrocytes[J]. Chinese Journal of Tissue Engineering Research, 2011, 15(7): 1169-1173. DOI: 10.3969/j.issn.1673-8225.2011.07.007 |
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| Authors: | Zhang Ping Liu Bin Cai Dao-zhang Zhong Zhi-hong Pan Yong-qian Zhang Zhen-shan |
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| Affiliation: | 1Department of Orthopaedics, the Third Affiliated Hospital of Guangzhou Medical University, Guangzhou 510150, Guangdong Province, China
2Department of Orthopaedics, the Third Affiliated Hospital of Sun Yat-sen University, Guangzhou 510630, Guangdong Province, China |
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| Abstract: | BACKGROUND:Interleukin-1 receptor antagonist (IL-1Ra) can delay osteoarthritis progress. The expression of IL-1Ra can be increased by gene transfection.OBJECTIVE:To construct recombinant human IL-1Ra protein fluorescent plasmid and to observe its expression in chondrocytes by liposome gene transfection. METHODS:pGEM-T-IL-1Ra was digested with double enzymes restriction, and then it was linked with T4 DNA ligase and cloned into the pEGFP-C1 vector. Rabbit articular chondrocytes were isolated and cultured in vitro. The plasmids carrying the IL-1Ra gene were transfected into chondrocytes. Then the expression of the transgene was observed under a fluorescence microscope and detected by using real-time PCR assay.RESULTS AND CONCLUSION:The sequence of obtained pEGFP-C1-IL-1Ra was identical to IL-1Ra and sequence in the Genbank. The expression of enhanced green fluorescent protein was observed. Real-time PCR analysis showed that the genes were expressed in chondrocytes. |
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