表达PEDV-S1蛋白的重组枯草芽孢杆菌构建及鉴定

Construction and Expression of the Recombinant PEDV-S1 in Bacillus subtilis

猪流行性腹泻病毒(PEDV)可引起新生仔猪严重腹泻和死亡,给养猪业造成巨大经济损失。根据枯草芽孢杆菌(Bacillus subtilis)调节动物胃肠道菌群平衡的功能和结构特征,选择其作为PEDV S1蛋白表达的宿主菌,构建重组基因工程枯草芽孢杆菌。实验将重组质粒p LA-PEDV-S1电转至枯草芽孢杆菌,提取枯草芽孢杆菌的重组质粒,采用PCR、Western blot和免疫荧光方法鉴定阳性质粒DNA。获得了重组pLA-PEDV-S1/B.Subtilis,S1蛋白成功展示表达在菌体表面,重组PEDV S1蛋白能够和猪源PEDV阳性血清结合,具有反应原性。该重组菌可能成为预防PEDV感染的口服疫苗候选菌株。

Porcine epidemic diarrhea(PEDV)couldcause piglets serous diarrhea and make them dead,which made huge economic losses to the pig industry.Based on the structure and specific characteristics of Bacillus subtilis for regulating the gastrointestinal tract flora balance in animal,the B.Subtiliswas chosen forthe host cell expressing S1 protein of PEDV.The recombinant pLAPEDV-S1/B.subtiliswas constructed.The extracted plasmid DNA were transferred to the competent cells B.subtilis.The positive recombinant bacteria were identified by PCR.Western Blot and immunofluorescence were employed to detect the expressed S1 protein.the recombinant B.Subtilis successfully surface-displayed the S1 protein.Western Blot showed that the expressed S1 protein could interact with anti-PEDV pig serum,suggesting it was reactionogenicity.thepLA-PEDV-S1/B.Subtilismight be a mucosal vaccine candidate for preventing piglets from PEDV infection.