表达猪白细胞介素2(pIL-2)重组乳酸杆菌的构建  

Construction of recombinant Lactobacillus expressing Pig interleukin 2(pIL-2)

作　　者：沙万里 闫满 刘仲藜 韦舒婉 李国江 SHA Wan-li;YAN Man;LIU Zhong-li;WEI Shu-wan;LI Guo-jiang(College of Animal Science and Technology,Jilin Agricultural Science and Technology University,Jilin 132101,China;Key Lab of Preventive Veterinary Medicine in Jilin Province,Jilin 132101,China)

机构地区：[1]吉林农业科技学院动物科技学院,吉林吉林132101 [2]预防兽医学吉林省重点实验室,吉林吉林132101

出　　处：《中国兽医杂志》2018年第8期37-41,共5页Chinese Journal of Veterinary Medicine

基　　金：吉林省教育厅项目(吉教科合字[2016]第206号);吉林农业科技学院大学生科技创新项目(吉农院合字[2014]第010号);(吉农院合字[2017]第047号)

摘　　要：为了建立表达猪白细胞介素2(pIL-2)的重组乳酸杆菌,用PCR的方法从pMD-18T-pIL-2质粒中扩增pIL-2基因,并送至吉林省库美生物科技有限公司进行测序分析。将测序正确的PCR扩增产物与pSIP409表达载体链接,通过PCR和双酶切方法鉴定重组质粒。将重组载体电转至乳酸杆菌,通过SDS-PAGE电泳和Western Blot的方法鉴定pIL-2蛋白的表达,通过细胞粘附实验检测重组乳酸杆菌对猪肠上皮细胞IPEC-J2细胞的粘附性。结果显示,通过PCR和酶切的方法鉴定pMD-18T-pIL-2质粒中基因大小与pIL-2基因大小相似,吉林省库美生物科技有限公司测序结果与Gen Bank上pIL-2基因100%吻合。通过SDS-PAGE电泳和Western Blot的方法鉴定,重组乳酸杆菌表达蛋白为pIL-2蛋白。重组乳酸菌可使猪肠道上皮细胞IPEC-J2结构紧密,细胞间粘连蛋白表达量增多。表明重组乳酸杆菌表达的蛋白为猪白细胞介素2(pIL-2)蛋白。To establish a recombinant Lactobacillus expressing porcine interleukin 2 （pIL - 2）. The pIL - 2 gene was amplifiedfrom pMD - 18T - pIL - 2 plasmid by PCR, and was sent to Jilin Ku Mei Biotechnology Co. , Ltd. for sequencing and analysis. Thecorrect PCR amplification products were linked with the pSIP409 expression vector, and the recombinant plasmid was identified byPCR and double enzyme digestion. The recombinant vector was transferred to Lactobacillus, and the expression of pIL - 2 protein wasidentified by SDS - PAGE electrophoresis and Western blot. The adhesion of recombinant Lactobacillus to porcine intestinal epithelialcells IPEC - J2 was detected by cell adhesion assay. The gene size of pMD - 18T - pIL - 2 plasmid was similar to that of pIL - 2gene by PCR and enzyme digestion. The sequencing results of pMD - 18T - pIL - 2 gene in Jilin province was the same as pIL - 2gene with a 100% sequence similarity. The expressed protein of the recombinant Lactobacillus was identified as pIL - 2 protein bySDS - PAGE electrophoresis and Western blot. Recombinant Lactobacillus increased the expression of IPEC - J2 and the intercellularadhesion protein in pig intestinal epithelial cells. The protein expressed by recombinant Lactobacillus is the protein of porcine inter-leukin 2 （pIL - 2）.

关 键 词：白细胞介素2 乳酸杆菌 表达 

分 类 号：S852.61[农业科学—基础兽医学]