重组野猪α-干扰素基因的高效表达和下游工艺的研发

利用PCR方法，从东北野猪和长白猪的肌肉基因组DNA中分别扩增出一条501bp的基因片断 ，与GenBank上登载的猪α-干扰素基因序列相比,核苷酸同源性达到98.4%以上。以其中野猪α-干扰素基因片断为参考模板，经过密码子优化、5?和3?区域的mRNA二级结构优化，全基因合成得到编码野猪α-干扰素基因成熟肽的基因。克隆该基因到原核表达载体pWL之中.随后转化至宿主菌DH-5α。经过SDS-PAGE电泳检测，发酵后的重组野猪IFN-α-干扰素基因的蛋白表达量大于25%。复性纯化后的重组蛋白，经过VSV-WISH系统检测，其生物活性为4.45×106IU/mg。

Overexpression And Down-Stream Technology of Recombinant Sus scrofa Interferon alpha in Escherichia coli.

A 501bp sequence of Sus scrofa IFN-αgene and Susscrota domestica of changbai IFN-αgene was amplified by normal polymerase chain reaction (PCR) from their genome whose homology are more than 98.4% ，compared with nucleotide sequence published on NCBI .Referring to Sus scrofa IFN-αgene sequence,a new sequence was synthesized after codon bias and mRNA structure optimizaed .Then ,this sequence was inserted in pWL vector and the recombinant plasmids were transformed into the E.Coli DH-5α.After fermentation,the expression level of recombinant protein are 25% by SDS-PAGE analysis.The biologic activity was 4.45×106 IU/mg by VSV-WISH system analysis after protein refolding and purification.