重组鸡IFN-α的高效表达与一步复性和纯化  

High expression and simultaneous renaturation and purification of the recombinant chicken IFN-α

作　　者：魏双施[1] 王艺萌[1] 王巍[1] 孙仰峰[1] 王君伟[1] 

机构地区：[1]东北农业大学动物医学学院,黑龙江哈尔滨150030

出　　处：《中国兽医科学》2010年第4期415-420,共6页Chinese Veterinary Science

摘　　要：通过在线网站以及生物学软件分析,将天然鸡IFN-α基因中的稀有密码子同义替换为大肠杆菌偏好的密码子。优化后的基因经人工合成后与温控型表达载体pWL连接并转入大肠杆菌中进行诱导表达,经SDS-PAGE分析,表达产物的分子质量约为19 ku,表达量占总蛋白的28.3%。表达产物主要以包涵体形式存在,用高浓度变性剂溶解后,采用Sephadex G50凝胶过滤一步复性并纯化重组鸡IFN-α,纯度可达98%以上。每1 L培养基可以得到47.1 mg的重组鸡IFN-α,产出率达34.14%。表达的重组鸡IFN-α在CEF/NDV检测系统上的比活性为1.12×108U/mg。The IFN-α gene of chicken was synthesized using optimized codons based on online analysis tools and bio-software and cloned into the pWL expression vector.The recombinant plasmid was transformed into Escherichia coli for expression.SDS-PAGE analysis showed that the expressed protein had a molecular weight of 19ku and accounted for 28.3% of the total cellular protein.The expression product mainly existed in the form of inclusion bodies.The dissolved inclusion body proteins were denatured and renatured simultaneously to purify the chicken IFN-α by gel filtration using Sephadex G50.The purity of the renatured IFN-α was exceed 98%and the yield reached 47.1mg/L（34.14%） and the resultant IFN-α showed a strong antiviral activity of 1.12×10^8U/mg on CEF/NDV culture.

关 键 词：鸡 Α-干扰素 基因优化 一步法 复性和纯化 

分 类 号：Q511[生物学—生物化学] Q786