大肠埃希菌K88ac菌毛蛋白亚基因的原核表达及其免疫原性研究

从E.coli C83902中扩增出不含信号肽序列的K88ac菌毛蛋白亚基基因片段,将其克隆到表达载体pQE-30中,构建了原核表达载体pQE30-K88ac,并转入E.coli XL1-Blue中,经IPTG诱导后,重组蛋白以包涵体形式获得高效表达.Western blot结果显示,表达的蛋白能够被K88ac单抗识别,用纯化的蛋白免疫小鼠,能够抵抗1 MLD大肠埃希菌强毒株C83902的攻击,这表明构建的工程菌株XL1-Blue(pQE30-K88ac)可以作为预防幼畜大肠埃希菌性腹泻基因工程菌苗的候选株.

Study on the Cloning and Expression of K88ac Fimbrial Subunit Gene and the Immunogenicity of Recombination Protein

The DNA fragment of K88ac fimbrial subunit gene without signal peptide sequence was amplified by PCR from the DNA of E.coli C83902 strain.After DNA cloning and sequencing,the fragment was inserted into expression vector pQE-30.The recombinant expression plasmid pQE30-K88ac was transferred into E.coli strain XL1-Blue,and the high level expressed recombinant protein was detected in the inclusion body protein of the expression strain induced by IPTG.The expressed protein can recognized by MAb of K88ac with western blot method.the mice which were immunized with the recombinant protein and then challenged with C83902 virulent strain had been protected.Therefore,the recombinant strain could be used as a candidate strain of gene engineering inactivated vaccine against colibacillus diarrhea of newborn piglet.