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源于草鱼肠道抗菌肽的重组表达、分离纯化和抗菌活性分析

发表时间:2012-06-15  浏览量:1146  下载量:354
全部作者: 史玉婷,孙科军,陶虚谷,苏建明,曹蓉,陈韬
作者单位: 湖南农业大学动物医学院;湖南农业大学动物科技学院
摘 要: 根据草鱼肠道中分离得到的抗菌肽(antibacterial peptide, AP)氨基酸测序结果,人工合成草鱼抗菌肽基因,并将该合成基因克隆至pET32a载体中,构建成含有6×His标签的重组质粒pET- AP. 重组质粒转化至大肠杆菌BL21中,经异丙基硫代半乳糖苷(IPTG)诱导后,进行十二烷基硫酸钠- 聚丙烯酰胺凝胶电泳(SDS- PAGE)分析。测序结果表明:插入片段的序列和位置均正确。通过Ni2+- NTA柱亲和层析纯化,用肠激酶将纯化的重组蛋白中的His标签切除后回收重组抗菌肽,采用滤纸片方法测定其抗菌活性。实验结果表明,重组抗菌肽能明显抑制大肠杆菌的生长。
关 键 词: 兽医学;抗菌肽;重组表达;纯化;抗菌活性
Title: Expression, purification and antimicrobial activity analysis of antimicrobial peptide originated from intestine of grass carp
Author: SHI Yuting, SUN Kejun, TAO Xugu, SU Jianming, CAO Rong, CHEN Tao
Organization: Veterinary Faculty, Hunan Agricultural University; College of Animal Science and Technology, Hunan Agricultural University
Abstract: According to the amino acid sequence of intestine of grass carp, antibacterial peptide (AP) coding genes were synthesized artificially and cloned into vector pET32a to construct a recombinant plasmid termed pET- AP which contains a 6譎is tag. Plasmid pET- AP was transferred to E.coli strain BL21. SDS- PAGE analysis of recombinant protein was carried out after induced by IPTG. Sequencing results indicated that the sequence and location of insert fragment in pET- AP were correct. Recombinant protein was purified by Ni2+- NTA column affinity chromatography. And enterokinase was used to recover the recombinant AP. Antimicrobial activity assay showed that recombinant AP could inhibit the growth of E.coli.
Key words: veterinary medicine; antibacterial peptide; recombinant expression; purification; antimicrobial activity
发表期数: 2012年6月第11期
引用格式: 史玉婷,孙科军,陶虚谷,等. 源于草鱼肠道抗菌肽的重组表达、分离纯化和抗菌活性分析[J]. 中国科技论文在线精品论文,2012,5(11):1081-1086.
 
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