摘要
烯醇化酶参与支原体糖酵解、细胞黏附及免疫调节等多种生命活动,对其开展功能研究有助于了解鸡毒支原体黏附及感染机制。本研究通过重叠PCR克隆获得鸡毒支原体烯醇化酶,经原核表达获得可溶性融合蛋白rMGEno,将纯化产物免疫BALB/c小鼠后制备获得特异性多抗。通过Western blot和ELISA检测发现,烯醇化酶在鸡毒支原体不同强弱毒株中高度保守,表达产量高,在鸡毒支原体参试各毒株的细胞膜中均有分布。这一表达产物的成功制备为深入研究鸡毒支原体烯醇化酶的生物学功能奠定了基础。
Enolase,known as an important enzyme involved in many life activities,such as glycolysis pathway,cell adhesion,and immunological regulation,is present in Mycoplasma gallisepticum（MG）.The study on biological function of MG enolase will be helpful to the understanding on the mechanisms of adherence and invasion.In the present study,MG enolase was amplified by overlapping PCR and subcloned into pET32a（＋） vector.With induction of IPTG,the soluble fusion protein,designated as rMGEno,was expressed in E.coli（DE3） and purified with His Resin Band kit.Polyclonal antibodies were prepared by immunizing three BALB/c mice using purified protein.Enolase was highly consistent and distributed on the membrane surface in virulent and attenuated MG strains as demonstrated in Western blot and ELISA.Further study will focus on the biological functions of MG enolase.
出处
《中国动物传染病学报》
CAS
2010年第3期26-31,共6页
Chinese Journal of Animal Infectious Diseases
基金
国家自然科学基金资助(30871883)
中央科研院所基础业务经费(2008JB12)
上海市科技兴农重点攻关项目(沪农科攻字[2007]第11-2号)
关键词
鸡毒支原体
烯醇化酶
可溶性表达
Mycoplasma gallisepticum
enolase
soluble expression
