猪型布鲁氏菌rOmp31_(48-74)-BLS融合肽的诱导表达与纯化

Induced Expression and Purification of rOmp31_(48-74)-BLS Fusion Peptide from Swine Brucella

目的:为了研究布鲁氏菌Omp31分子的抗原性质,以及布鲁氏菌BLS蛋白质的聚合功能,通过融合PCR技术,制备布鲁氏菌Omp3148-74与BLS的融合蛋白质,并且进行蛋白质的纯化。方法:以猪型布鲁氏菌Omp31基因和BLS基因作为研究对象,通过融合PCR技术构建重组表达载体,IPTG诱导融合蛋白质表达之后,利用His-tag亲和层析柱进行亲和纯化。结果:Omp3148-74-BLS融合DNA长度为531 bp,编码177个氨基酸;加上原核表达载体上的His-tag标签,融合蛋白质的实际大小为25.07 kD,实验结果证实表达载体构建正确,重组表达正确。结论:通过融合PCR技术构建的重组子pET-28-a-Omp3148-74-BLS,可以在大肠杆菌中成功表达;His-tag亲和层析技术可以纯化到Omp3148-74-BLS融合蛋白,而且蛋白质纯度较高。

Objective:To study the antigenic propertity of Brucella Omp31 molecules,as well as the aggregation function of BLS protein,the integration PCR technology was used to prepare the Omp3148-74-BLS fusion protein.And purification of this fusion protein was done.Method:Using swine Omp31 gene and BLS gene as the research object,recombinant expression vector was constructed through integration PCR technology.After inducing by IPTG,the fusion protein was purified by His-tag affinity chromatography.Result:Omp3148-74-BLS fusion DNA was made up of 531 bp,encoding 177 amino acids.After adding the His-tag label from prokaryotic expression vector,the actual size of fusion protein was 25.07kD.And experimental results confirmed that expression vector was correctly constructed.Conclusion:The pET-28-a-Omp3148-74-BLS recombinant expression vector that constructed through integration PCR technology could be successfully expressed in Escherichia coli.And Omp3148-74-BLS fusion protein could be purified by His-tag affinity chromatography.The protein purity was much higher.