摘要
为建立生物合成牛乳铁素的方法,根据牛乳铁素氨基酸序列确定其编码序列,利用PCR技术获得基因扩增产物,接着将该产物与编码鱼腥蓝细菌Anabaena sp.PCC7120的DNA聚合酶基因dnaENI中断裂的内含肽基因Asp dnaEIn的PCR产物进行重组PCR,将获得的重组融合基因再克隆到表达载体pET-28a构建重组融合表达质粒。结果显示,融合表达质粒转化大肠杆菌后经诱导能在大肠杆菌表达宿主Escherichia.coli BL21(DE3)中大量表达,且细胞裂解液与纯化的Anabaena sp.PCC7120中含断裂内含肽Asp DnaEIc的DNA聚合酶蛋白DnaECI混合反应后对金黄色葡萄球菌具有抑制作用。
In order to establish a bovine lactoferricin biosynthesis method,the bovine lactoferricin-encoding gene was determined on the basis of amino acid sequence. A recombinant fusion gene was obtained by means of overlapping PCR following the purification of the separate PCR products of the bovine lactoferricin-encoding sequence and the split intein Asp dnaEIn gene from the DNA polymerase gene dnaENI of Anabaena sp. PCC 7120,and the obtained fusion gene was subsequently cloned into the expression plasmid pET-28a to construct a recombinant fusion expression plasmid. The results showed that recombinant fusion proteins were over-expressed after the expression host Escherichia coli BL21 （DE3） containing the recombinant bovine lactoferricin fusion expression plasmid was induced and that reaction products of over-expressed recombinant bovine lactoferricin lysates and the purified Anabaena sp. PCC 7120 DnaECI,which contained split intein Asp DnaEIc,could inhibit Staphylococcus aureus.
出处
《食品科学》
EI
CAS
CSCD
北大核心
2010年第19期295-298,共4页
Food Science
基金
高等学校博士点科研基金项目(20090204120028)
陕西省自然科学基金项目(SJ08B12)
关键词
牛乳铁素
重组PCR
融合基因
融合蛋白
bovine lactoferricin
overlapping PCR
fusion gene
fusion protein
