Zhang Dongmei, Pan Weiqing, Lu Deru, Jiang Liping
Institute of Medical Biotechnology & Molecular Genetics of Second Military Medical University, Shanghai 200433 China.
Zhonghua Yi Xue Za Zhi. 2002 Feb 10;82(3):198-202.
Production of 3D7/MSP1 - 42 recombinant protein with correct conformation in Pichia pastoris for vaccine efficiency assay.
Asymmetric PCR-based method was utilized to synthesize the 1 202 bp 3D7/msp1 - 42 gene. The expressing plasmid containing the synthetic gene was introduced into Pichia pastoris by electroporation. The secreted product was detected by Western Blot.
The redesigned entire 3D7/msp1 - 42 gene was generated with error-free, and expressed to produce 42 kD recombinant protein in secreted form. Conformational monoclonal antibody specific for MSP1 C-terminal can interact with the recombinant protein.
The redesigned 3D7/msp1 - 42 gene was expressed in P. pastoris with full length of recombinant protein which resembled most likely to the native protein.
在毕赤酵母中生产具有正确构象的3D7/MSP1 - 42重组蛋白用于疫苗效率检测。
采用基于不对称PCR的方法合成1202 bp的3D7/msp1 - 42基因。通过电穿孔将含有合成基因的表达质粒导入毕赤酵母。用蛋白质免疫印迹法检测分泌产物。
重新设计的完整3D7/msp1 - 42基因无错误生成，并以分泌形式表达产生42 kD重组蛋白。对MSP1 C末端具有特异性的构象单克隆抗体可与重组蛋白相互作用。
重新设计的3D7/msp1 - 42基因在毕赤酵母中表达，产生的重组蛋白全长与天然蛋白极为相似。
