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目的 :对人IFNα_2b基因的 5′端进行改造 ,从翻译水平上调控IFNα_2b基因在大肠杆菌中的表达。方法 :采用PCR方法 ,人工合成寡核苷酸引物 ,对人IFNα_2b基因进行了改造 :去除3′端非编码区 ,并增加原核系统强终止密码子。将改造后的基因片段分别克隆入原核表达载体pBV32 1,在大肠杆菌中进行表达。对表达产物进行活性效价测定。结果 :3′端删除非编码区 ,表达水平比删除前提高 4倍。结论 :IFNα_2b基因的 3′端非编码区对其在原核系统的表达具有抑制作用 ,删除该区可提高表达水平。
OBJECTIVE: To modify the 5 ’end of human IFNα_2b gene and regulate the expression of IFNα_2b gene in Escherichia coli at translation level. Methods: Human IFNα_2b gene was modified by PCR and oligonucleotide primers were synthesized. The 3 ’noncoding region was removed and the strong stop codon of prokaryotic system was added. The modified gene fragments were cloned into the prokaryotic expression vector pBV32 1, and expressed in E. coli. The expressed product was assayed for activity titer. Results: The non-coding region was deleted at the 3 ’end, and the expression level was increased by 4 times compared with that before deletion. Conclusion: The 3 ’untranslated region of IFNα_2b gene can inhibit the expression of IFNα_2b gene in prokaryotic system. The deletion of this region can increase the expression of IFNα_2b gene.