摘要
本研究用PCR方法从停乳链球菌C588的基因组DNA中扩增出MIG基因,用T/A克隆法将其插入pBS-T载体,并构建原核表达载体pET-32a(+)-MIG。用BL21(DE3)/pET系统表达Trix-MIG融合蛋白,SDS-PAGE和Westernblot分析鉴定表达产物。结果PCR扩增产物经测序,证实与GenBank中停乳链球菌MIG基因(AF354651)序列同源性为99%。SDS-PAGE显示,经IPTG诱导后BL21(DE3)/pET-32a(+)-MIG总蛋白中出现一条相对分子质量为89ku的新蛋白条带。Westernblot分析显示,MIG蛋白可与停乳链球菌多克隆抗血清发生特异性反应。MIG融合蛋白的表达为MIG在细菌致病中的作用研究以及相关疫苗的制备奠定了基础。
To express the gene encoding MIG of streptococcus dysgalactiae and provide target protein for further immunological study, MIG gene from the genomic DNA of standard streptococcus dysgalactiae strain C588 by was amplified by PCR and insert into pET-32a （＋） vector. The recombinant vector pET-32a （＋）-MIG was transformed into BL21 （DE3）/pET system for expression of Trix-MIG fusion protein. The expressed product was identified by SDS-PAGE and Western blot. The result showed that the MIG gene shared 99 % sequence identity with that of Streptococcus dysgalactiae （AF354651） reported in GenBank with the protein had a relative molecular weight of 89 ku, and could be recognized by specific antibodies in Western blot.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2008年第6期430-434,共5页
Chinese Journal of Preventive Veterinary Medicine
关键词
停乳链球菌
MIG
克隆与表达
Streptococcus dysgalactiae
MIG
cloning and expression
