Position: Home > Articles > Prokaryotic Expression and Purification of Canine β-defensin 103
Progress in Veterinary Medicine
2017,38
(5)
26-29
犬β-防御素cBDl03的原核表达和纯化
作 者：
朱骞;包银莉;秦海斌;宋珍华;贺星亮;温海;高一龙
关键词：
犬阻防御素103;原核表达;纯化
摘 要：
将犬β-防御素103(canine β-defensin103,cBD103)基因亚克隆至原核表达栽体pET-32a(+),构建重组表达质粒pET-cBD103,转化大肠埃希菌BL21感受态细胞,IPTG诱导表达后,利用Ni-NTA亲和层析纯化该蛋白,并用肠激酶酶切融合蛋白,SDS-PAGE鉴定表达产物和纯化产物.结果表明,质粒pET-cBD103经PCR及双酶切鉴定证明构建正确,大肠埃希菌中成功表达出目的蛋白,蛋白大小约为24ku,以可溶性表达为主.经过纯化和酶切,得到大小8ku的cBD103,与预期大小一致.该研究在大肠埃希菌中表达了目的蛋白,为下一步大规模制备cBD103奠定了基础.
译 名：
Prokaryotic Expression and Purification of Canine β-defensin 103
关键词：
canine β-defensin 103%prokanyotic expression%purification
摘 要：
The canine β-defensin 103 (cBD103) gene was subcloned to pET-32a(+) vector,and the con- structed recombinant plasmid pET-cBD103 was transformed to competent E.coli BL21 for expression under induction of IPTG.The expressed product was purified by nickel ion affinity chromatography and iden- tified by SDS-PAGE. Results of both PCR and enzyme digestion analysis proved that recombinant pET-cBD103 was constructed correctly.The expressed protein,with a relative molecular mass of 24 ku, mainly existed in a soluble form.After purification and restriction enzyme digestion,a protein band about 8 ku was detected, and consistent with the purpose bands.The cBD103 protein was successfully expressed in E.coli, which laid a foundation for further preparation of cBD103 protein and clinical test.
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