| 人中性粒细胞多肽1,3真核表达载体的构建与鉴定 |
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| 引用本文： | 刘娟,孙永涛,杜德伟,李光玉,刘秋平,翟嵩.人中性粒细胞多肽1,3真核表达载体的构建与鉴定[J].医学研究生学报,2004,17(10):868-871. |
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| 作者姓名： | 刘娟 孙永涛 杜德伟 李光玉 刘秋平 翟嵩 |
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| 作者单位： | 第四军医大学唐都医院,解放军感染病诊疗中心,陕西西安,710038 |
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| 摘 要： | 目的：扩增人中性粒细胞多肽1，3(HNP1，3)基因片段，构建真核表达载体pcDNA3．1／V5-His-TOPO／HNP1，3。方法：从健康人中性粒细胞中提取总RNA，用RT-PCR方法扩增出HNP1，3基因完整的cDNA片段，应用TA克隆插入pMD18-T载体，经酶切鉴定及序列分析确证后，将其亚克隆至真核表达载体pcDNA3．1／V5-His-TOPO中。结果：从人中性粒细胞中克隆出人HNPl1，3基因，经测序分析与GenBank公布的人HNP1，3核苷酸序列同源性为100％，成功地构建了真核表达载体pcDNA3．1／V5-His-TOPO／HNP1，3。结论：真核表达载体pcDNA3．1／V5-His-TOPO／HNP1，3的成功构建，为后续重组基因的真核／表达及其对HIV-1抑制作用的研究奠定了实验基础。
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| 关 键 词： | 中性粒细胞多肽 反转录-聚合酶联反应 基因 克隆 |
| 文章编号： | 1008-8199(2004)10-0868-04 |
| 修稿时间： | 2004年5月18日 |
Construction and identification of human neutrophil peptide eukaryotic expression vector |
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| LIU Juan,SUN Yong-tao,DU De-wei,LI Guang-yu,LIU Qiu-ping,ZHAI Song.Construction and identification of human neutrophil peptide eukaryotic expression vector[J].Bulletin of Medical Postgraduate,2004,17(10):868-871. |
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| Authors: | LIU Juan SUN Yong-tao DU De-wei LI Guang-yu LIU Qiu-ping ZHAI Song |
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| Abstract: | Objective:To obtain encoding gene segment of human neutrophil peptides (HNP1,3) and construct the eukaryotic expression plasmid pcDNA3.1/V5-His-TOPO- HNP1,3. Methods:RNA was extracted from human neutrophil cell as template, cDNA encoding mature HNP1,3 was amplified by RT-PCR and inserted into cloning vector pMD18-T. After restriction endonuclease digestion and DNA sequencing confirmation the gene was subcloned into eukaryotic expression plasmid pcDNA3.1/V5-His-TOPO. Results:The gene encoding HNP1,3 was cloned whose DNA sequence was 100% identical with that published in GenBank, a recombinant expression plasmid pcDNA3.1/V5-His-TOPO/HNP1,3 was successfully constructed. Conclusion:The successful construction of pcDNA3.1/V5-His- TOPO/HNP1,3 lies a basis for eukaryotic expression and further research on anti-HIV activity of HNP1,3. |
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| Keywords: | Human neutrophil peptides RT-PCR Gene Clone |
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