牛β-防御素BNBD11基因在大肠杆菌中的表达

Expression of Bovine Neutrophil β-Defensin BNBD11 Gene in Escherichia coli

探索牛β-防御素BNBD11原核表达及纯化的技术路线。根据已报道的BNBD11的氨基酸序列,兼顾大肠杆菌密码子偏好性,设计合成BNBD11基因。构建重组表达载体pET32a-BNBD11,转入E.coli BL21,用异丙基-β-D-硫代吡喃半乳糖苷(IPTG)诱导表达。经SDS-PAGE检测融合蛋白结果显示,大部分表达产物以可溶形式存在,用Ni Sepharose柱层析纯化融合蛋白。融合蛋白经甲酸切割后,再次用Ni Sepharose柱层析去除带有His-Tag的杂蛋白,最终得到纯化的重组BNBD11。实验实现了BNBD11在大肠杆菌中的融合表达,并纯化了获得的重组BNBD11。

In order to explore the technical route for prokaryotic expression and purification of bovine neutrophilβ-defensin 11（BNBD11）,BNBD11 gene was synthesized using preferred condons of E.coli,according to the reported amino acid sequence of BNBD11.The recombinant expression plasmid pET32a-BNBD11 was constructed and then transformed into E.coli BL21.The expression of BNBD11 in E.coli BL21 was induced by isopropylβ-D-1-thiogalactopyranoside（IPTG）.SDS-PAGE showed that most expressed products were soluble.The fusion protein was purified by Ni Sepharose column chromatography.After the cleavage of the fusion protein by formic acid,Ni Sepharose column was used again to remove proteins with His-Tag and the purified recombinant BNBD11 was achieved.Therefore,BNBD11 was successfully expressed in E.coli,and the recombinant BNBD11 was obtained.