摘要
利用含有强启动子 PAOX1 和 α-因子信号肽序列的巴斯德毕赤酵母载体 p PICZαA,构建出含牛白细胞介素 2(Bo IL - 2 )基因的重组质粒 Bo IL 2 - p PICZαA。线性化的重组表达载体转化到巴斯德毕赤酵母 X- 33及 KM71H中 ,筛选Zeocin高抗性酵母菌株 ,甲醇诱导目的蛋白表达。经 SDS- PAGE和 Western blot检测表明 ,Bo IL- 2以融合蛋白形式在胞内表达 ,但没能分泌到胞外。通过 Bo IL- 2在巴斯德毕赤酵母中的表达 ,重点讨论了信号肽。
The bovine Interleukin-2 gene was inserted into the Pichia pastoris expression vector pPICZαA which contains AOXI promoter and α-factor signal sequence.The recombinant plasmid of BoIL-2pPICZαA was linearnized by SacⅠ and transformed into X-33 and KM71H by electroporation.The recombinant Pichia strains.identified by PCR and Zeocin resistance were cultured and induced with methanol.SDS-PAGE analysis and Western blot showed BoIL-2 was expressed in a fused protein with the α-factor signal peptide but failed to secrete into medium.Effect of signal peptide and prefer codon on efficiency of secretory expression was also discussed in detail.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2004年第3期261-263,267,共4页
Chinese Journal of Veterinary Science
基金
上海市科技兴农重点攻关项目 (科农攻字 2 0 0 1第 3 -7号 )
