柔嫩艾美耳球虫LZ株MIC-5基因的克隆与表达  

Cloning sequencing and expression of microneme protein 5 gene of Eimeria tenalla LZ strain

作　　者：孟庆玲[1] 乔军[1] 才学鹏[1] 景志忠[1] 田广孚[1] 闫鸿斌[1] 

机构地区：[1]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室甘肃省动物寄生虫病重点实验室

出　　处：《甘肃农业大学学报》2007年第6期8-12,共5页Journal of Gansu Agricultural University

基　　金：国家重大基础研究"973"项目(No.G1999011906)

摘　　要：提取柔嫩艾美耳球虫兰州株孢子化卵囊总RNA，运用RT-PCR技术扩增＆MIC-5基因片段进行测序，并克隆入大肠杆菌表达载体pET28a中进行诱导表达．结果表明，该基因片段长918bp，编码306个氨基酸．与GenBank报道的柔嫩艾美耳球虫Houghton株MIC-5基因相比，核苷酸和推导的氨基酸序列同源性为99．2％和99．6％．在推导的MIC-5氨基酸序列中，第15～89、90～160、173～242和245～306位保守的cys残基形成4个Apple样结构域（A-domain）．推测该蛋白可能在虫体与宿主细胞的黏附中起重要作用．转化重组质粒pETMIC-5的BL21（DE3）经IPTG诱导后，SDS—PAGE和western blot分析证实目的蛋白成功表达，重组蛋白的表达量可占菌体蛋白的15．2％．Microneme protein 5 gene of Eimeria tenalla LZ strain was amplified by RT-PCR and cloned into pMD18-T for sequencing, then subcloned into the prokaryotic expressing vector pET28a for expression. The amplified fragment length of Et MIC-5 gene was 918 bp, encoding 306 amino acids. Compared with the MIC-5 genes of Houghton strain, the identities of nucleotide sequence and deduced amino acid sequence were 99.2 % and 99. 6 %,respectively. There were four Apple domains （15-89,90- 160,173-242 and 245-306） in the deduced amino acid sequence,which implied that MIC,5 protein may play roles in invasion of Eimeria tenalla. Moreover,E. coli strain BL21 （DE3） transformed with recombinant pETMIC--5 can express interesting protein, which amounts to 15.2 % in the total protein of the induced bacteria by the assaying of gel scanning.

关 键 词：柔嫩艾美耳球虫 微线蛋白5基因 克隆 表达 

分 类 号：S851.347.203[农业科学—预防兽医学] Q786[农业科学—兽医学]