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HNP1,3基因 ,经测序分析与GenBank公布的人HNP1,3核苷酸序列同源性为 10 0 % ,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
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.
出处
《医学研究生学报》
CAS
2004年第10期868-871,共4页
Journal of Medical Postgraduates
