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[猪瘟病毒T细胞表位E290肽与猪细小病毒VP2蛋白在干酪乳杆菌中的共表达及免疫小鼠体内特异性抗体的测定]

[Co-expression of CSFV T cell epitope E290 peptide and PPV VP2 protein in Lactobacillus casei and determination of specific antibodies in immunized mice].

作者信息

Xu Yi-gang, Cui Li-chun, Ge Jun-wei, Zhao Li-li, Li Yi-jing

机构信息

Department of Veterinary Medicine, Northeast Agriculture University, Harbin 150030, China.

出版信息

Wei Sheng Wu Xue Bao. 2007 Aug;47(4):667-72.

Abstract

Lactobacillus casei strain 393 was selected as an antigen delivery vehicle for the development of oral vaccine to express recombinant classical swine fever virus (CSFV) T cell epitope E290 peptide and porcine parvovirus (PPV) VP2 protein. The recombinant genes encoding CSFV T cell epitope E290 peptide and PPV VP2 protein, respectively, were cloned into the secretion expression vector pPG, and then the pPG-VP2-E290 was electrotransformed into L. casei 393 giving rise to recombinant strain pPG-VP2-E290/L. casei 393. The recombinant L. casei 393 was induced by 2% lactose in MRS and about 70kDa protein was detected with SDS-PAGE in induced recombinant strain and culture supernatants. The result of Western blot indicated that the expressed protein possessed the antigenic specificity same as the native virus protein. The indirect ELISA test also indicated that the interest protein was expressed and secreted from the recombinant strain. Specific anti-PPV VP2 secret immunoglobulin A (sIgA) antibody was detected by indirect ELISA in the feces, anti-PPV VP2 and anti-CSFV E290 peptide immunoglobulin G (IgG) antibody was detected by indirect ELISA in the serum of immunized mice after intragastric administration. The results indicated that the mice immunized with recombinant strain pPG-VP2-E290/L. casei 393 could produce clear antibody level, which establish important material basement for the development of lactic acid bacteria oral vaccine of recombinant CSFV and PPV.

摘要

干酪乳杆菌菌株393被选作抗原递送载体用于开发口服疫苗,以表达重组经典猪瘟病毒(CSFV)T细胞表位E290肽和猪细小病毒(PPV)VP2蛋白。分别编码CSFV T细胞表位E290肽和PPV VP2蛋白的重组基因被克隆到分泌表达载体pPG中,然后将pPG-VP2-E290电转化到干酪乳杆菌393中,产生重组菌株pPG-VP2-E290/干酪乳杆菌393。重组干酪乳杆菌393在MRS中用2%乳糖诱导,诱导后的重组菌株和培养上清液经SDS-PAGE检测到约70kDa的蛋白。Western印迹结果表明,表达的蛋白具有与天然病毒蛋白相同的抗原特异性。间接ELISA试验也表明,目的蛋白是从重组菌株中表达和分泌的。经胃内给药后,通过间接ELISA在粪便中检测到特异性抗PPV VP2分泌型免疫球蛋白A(sIgA)抗体,在免疫小鼠血清中通过间接ELISA检测到抗PPV VP2和抗CSFV E290肽免疫球蛋白G(IgG)抗体。结果表明,用重组菌株pPG-VP2-E290/干酪乳杆菌393免疫的小鼠能够产生明显的抗体水平,这为重组CSFV和PPV乳酸菌口服疫苗的开发奠定了重要的物质基础。

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