牛IgG Fc片段与FnBPB-ClfA串联表达及其对金黄色葡萄菌性奶牛乳腺炎的免疫效果

发布时间：2018-05-21 20:42

本文选题：金黄色葡萄球菌 + 奶牛IgG　；参考：《动物医学进展》2017年10期

【摘要】：为了提高金黄色葡萄球菌(S.aureus)黏附素FnBPB-ClfA基因表达蛋白对奶牛的免疫效力,应用RT-PCR扩增奶牛IgG Fc基因片段与构建的金黄色葡萄球菌(S.aureus)FnBPB-ClfA黏附素基因串联构建质粒pMD19T-Fc-FnBPB-ClfA,并通过双酶切将其克隆于pET32a(+),以构建质粒pET32a-Fc-FnBPB-ClfA并进行诱导表达。以盐酸左旋咪唑(LH)或氢氧化铝(ALUM)为佐剂,与纯化的重组蛋白(bFcFnBPB-ClfA)混合以首次免疫肌肉注射,加强免疫乳头管灌注或首次免疫、二次免疫及三次免疫均以肌肉注射方式免疫临近干奶期的奶牛。用间接ELISA和Western blot对各组牛血清中的抗体进行监测,评价免疫效果。结果显示,以LH为佐剂免疫的(A组)牛21d时抗体水平最高,可达到1∶1 600;而以ALUM为佐剂免疫的B组和C组抗体水平分别为1∶800和1∶400。IgG水平动态监测结果表明,以LH为佐剂免疫牛的血清中特异性抗体上升趋势明显高于其他免疫组。研究结果表明,以首次免疫肌肉注射,加强免疫乳头管灌注的方式较好,并且bFc-FnBPB-ClfA与LH佐剂混合后免疫奶牛能最有效激发奶牛免疫反应。
[Abstract]:In order to improve the immune effect of the gene expression protein of Staphylococcus aureus (S.aureus) FnBPB-ClfA gene, the IgG Fc gene fragment was amplified by RT-PCR to construct the plasmid pMD19T-Fc-FnBPB-ClfA in series with the constructed Staphylococcus aureus (S.aureus) FnBPB-ClfA adhesion gene, and was cloned to pET32a (+) by double enzyme digestion. The plasmid pET32a-Fc-FnBPB-ClfA was constructed and induced to induce the expression. Using LH or ALUM as the adjuvant, the recombinant protein (bFcFnBPB-ClfA) was mixed with the purified recombinant protein (bFcFnBPB-ClfA) for the first immunization of the intramuscular injection. The immunization of the nipple tube was enhanced or the first immunization was enhanced. The two and three immunizations were immunized near the dry milk by intramuscular injection. The bovine serum antibody in each group was monitored with indirect ELISA and Western blot to evaluate the immune effect. The results showed that the antibody level of bovine 21d with LH as adjuvant was highest and reached 1: 1600, while the level of antibody water in B and C groups with ALUM as adjuvant immunity was 1 to 800 and 1: 400.IgG level dynamic monitoring results. The results showed that the increasing trend of specific antibodies in the serum of cattle immunized with LH was obviously higher than that of other immune groups. The results showed that the method of strengthening the immunization with the immunization of the nipple tube was better by the first immunization of the muscle, and the immunization of cows could be most effectively stimulated by the mixture of bFc-FnBPB-ClfA and LH adjuvant.
【作者单位】：内蒙古农业大学兽医学院微生物学与免疫学实验室;
【基金】：国家自然科学基金项目(31460664) 农牧交错区生态型现代奶业生产模式研究与示范项目(2012BAD12B09-02)
【分类号】：S858.23

【相似文献】