| 猪白细胞介素18基因的克隆及真核表达重组质粒的构建 |
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| 引用本文： | 石昂,时庆贺,李新果,王玉国,杨利,李双双,陈陆,王川庆.猪白细胞介素18基因的克隆及真核表达重组质粒的构建[J].中国畜牧兽医,2017,44(9):2587-2592. |
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| 作者姓名： | 石昂 时庆贺 李新果 王玉国 杨利 李双双 陈陆 王川庆 |
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| 作者单位： | 河南农业大学牧医工程学院, 郑州 450002 |
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| 基金项目： | 国家自然科学基金（31272567、31772781）；河南省高校科技创新人才支持计划（14HASTIT022） |
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| 摘 要： | 为获得表达猪白细胞介素18（interleukin-18，IL-18）的真核表达重组质粒，试验通过RT-PCR从猪脾脏、肺脏和淋巴结组织中扩增猪IL-18全基因并定向克隆到真核表达载体pZJ-1，测序分析和酶切鉴定正确后，转染至293T细胞中，并通过实时荧光定量PCR和Western blotting分别在基因和蛋白水平检测IL-18的表达。结果表明，试验成功构建了真核表达重组质粒pZJ-IL-18，且可以在293T细胞中表达IL-18基因。Western blotting试验证实，猪IL-18多克隆抗体能与约17 ku的表达产物发生特异性反应，表明IL-18能正确表达且具有反应原性。本试验构建了表达猪IL-18基因的真核表达质粒，并在293T细胞中瞬时表达，为进一步研究IL-18的功能奠定基础。
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| 关 键 词： | 猪 IL-18基因 真核表达载体 克隆 |
| 收稿时间： | 2017-05-24 |
Cloning and Eukaryotic Expression Plasmid Construction of Porcine Interleukin-18 Gene |
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| SHI Ang,SHI Qing-he,LI Xin-guo,WANG Yu-guo,YANG Li,LI Shuang-shuang,CHEN Lu,WANG Chuan-qing.Cloning and Eukaryotic Expression Plasmid Construction of Porcine Interleukin-18 Gene[J].China Animal Husbandry & Veterinary Medicine,2017,44(9):2587-2592. |
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| Authors: | SHI Ang SHI Qing-he LI Xin-guo WANG Yu-guo YANG Li LI Shuang-shuang CHEN Lu WANG Chuan-qing |
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| Institution: | College of Animal Husbandry and Veterinary, Henan Agricultural University, Zhengzhou 450002, China |
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| Abstract: | To obtain recombinant eukaryotic expression plasmid of porcine interleukin-18(IL-18), the whole gene of porcine IL-18 gene was amplified from porcine spleen, lung and lymph nodes by RT-PCR and cloned into eukaryotic expression vector pZJ-1. The recombinant expression plasmid pZJ-IL-18 were identified by enzyme digestion and sequencing analysis, and was transfected into 293T cells.The expression of IL-18 was detected by Real-time PCR and Western blotting in both gene and protein levels. The results showed that the eukaryotic expression plasmid of porcine IL-18 was constructed and could express transiently in 293T cells. Western blotting result confirmed that porcine IL-18 polyclonal antibody could react specifically with approximately 17 ku expression products,and indicated that IL-18 could express correctly and be responsive.This study constructed the eukaryotic expression plasmid of porcine IL-18 gene which could express transiently in 293T cells, and laid the foundation for studying function of IL-18. |
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| Keywords: | porcine IL-18 gene eukaryotic expression vector clone |
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