【摘要】 利用PCR技术从质粒pcDNA3.1/GS扩增得到hIL10基因.将其插入含有AOX1启动子和α分泌信号肽序列的毕赤酵母表达载体pPIC9K中,构建重组质粒pPIC9K/IL10,转化毕赤酵母GS115,筛选出整合了多拷贝白细胞介素10基因的菌株,经摇瓶培养,0.5%甲醇诱导表达.SDS PAGE分析显示,表达产物以可溶性分子形式存在于上清中,诱导4d的表达量达到高峰.Western印迹表明,表达产物具有良好的抗原性和特异性.更多还原

【Abstract】 The gene encoding human interleukin 10 was amplified with PCR from the pcDNA/3.1 plasmid, and then was cloned into the yeast expression vector pPIC9K containing the AOX1 promoter and the secretion signal sequence coding αfactor prepro-leader peptide. Then the yeast expression vector pPIC9K/hIL10 was constructed and transformed to P. pastoris GS115 strains, screened for multiple inserts, induced with0.5% methanol in shake flask cultures. Increasing level of rhIL10 was detected in the medium for up to 4 days by SDS-PAGE analysis. Western blot shows that the expressed rhIL10 exhibits high specificity and antigenicity.更多还原