鸡β-防御素7的克隆及表达  

Cloning and expression of avian beta defensin 7 gene from chicken

作　　者：易道生[1] 魏晓东 缪永建 陈燕飞[1] YI Dao-sheng;WEI Xiao-dong;MIAO Yong-jian;CHEN Yan-fei(Henry Fok College of Life Science,Shaoguan University,Shaoguan 512005,China)

机构地区：[1]韶关学院英东生命科学学院,广东韶关512005

出　　处：《广东农业科学》2018年第2期130-134,共5页Guangdong Agricultural Sciences

基　　金：韶关市科技计划项目(405-99000313);国家大学生创新创业项目(201410576007);韶关学院校级课题(314-140642)

摘　　要：以NCBI已登录的鸡β-防御素7(登录号:NM_001001194)为模板,设计特异性引物,扩增鸡β-防御素7基因,目的基因与pET32a载体相连,构建表达质粒并导入大肠杆菌BL21,经IPTG诱导表达和抗菌试验。结果表明,PCR扩增的目的片段,经测序证明与鸡β-防御素7基因NM_001001194序列同源性达99%,经IPTG诱导表达的重组蛋白与预期大小相符。平板抑菌试验结果表明,该重组多肽对金黄色葡萄球菌有较强的抗菌活性。A pairs of primers used in polymerase chain reaction(PCR)was designed following the NM_001001194 from GenBank to amplify the beta defensin 7 gene of chicken.The total RNA was isolated from spinal cord cells of chicken,and the first cDNA strand was obtained by approach of reverse transcription PCR.The products of PCR were linked with T plasmid vectors,and then transformed into Escherichia coli(E.Coli)DH5αstrain cells subsequently.Expressional plasmid vector,pET32a-gal7,was reconstructed with restriction endonuclease HindⅢand BamH I.Recombinant plasmid vector,pET32a-gal7,was induced with Isopropyl-β-D-Thiogalacto Pyranoside(IPTG)in E.Coli DE3 strain cells to produce recombinant Gal7 protein.The results showed that the cloned cDNA sequence of beta defensin 7 gene of chicken was comprised of 132 nucleotide acid residues,and encoded the beta defensin 7 with 44 amino acids and predicted molecular weight 4923.81Da.The identity to NM_001001194 is 99%.The recombinant peptides had strong antibacterial activity against staphylococcus aureus in vitro.

关 键 词：Β-防御素 克隆 原核表达 基因重组 

分 类 号：S813.3[农业科学—畜牧学] Q786[农业科学—畜牧兽医]