摘要
根据Genbank的犬干扰素α序列,经密码子优化后,设计并合成了编码CaIFN-α蛋白的基因片段。CaIFN-α基因与pBV220温度诱导表达载体连接,转化大肠杆菌BL21。经培养、升温诱导表达,目的蛋白以包涵体的形式存在,蛋白分子量约为18000,蛋白表达量为25%。菌体经超声破碎、变性、复性后,目的蛋白纯度为69%,收率为28%;疏水层析后,目的蛋白纯度87%,收率为81%;分子筛层析后,目的蛋白纯度为96%,收率为90%。纯化的CaIFN-α蛋白的生物学比活性为3×107 IU/mg,内毒素≤10 EU/mg,符合兽用药要求。此制备工艺总产率高达20%、重现性好,适用于大规模生产。
After codon optimization, the CalFN - a gene fragments was synthesized, cloned into pBV220 vector and expressed in E. coil BI21. The molecular weight of CalFN - α-was 18000 and the expression of CalFN - α was 25%. Then the inclusion denatured and renaturated, the purity was 69%, and recovery rate reached 28%. Hydrophobic interaction chromatography was used to purify the protein,the purity was 87%, and recovery rate reached 81%. Then the protein was purified by molecular sieving chromatography, the purity was 96%, and recovery rate reached 90%. The specific activity of CalFN - α was 3 x 107 IU/mg, the endotoxin was equal to or less than 10 EU/mg, and yield was 20%. This process for production was suitable for large -scale production.
出处
《中国兽药杂志》
2012年第3期44-47,共4页
Chinese Journal of Veterinary Drug
关键词
犬干扰素α
原核表达
纯化
质量
canine interferon α
prokaryotic expression
purification
quality
