摘要
借助计算机辅助设计了一种新的人降钙素类似物 (新型降钙素 )。根据密码子偏爱性原理及基因工程操作原则设计了它的基因 ,运用化学法和酶法相结合的技术合成了该基因 ,并克隆到质粒 pUC19中 ,DNA测序验证正确。该基因被克隆到融合型表达载体pGEX 2T中 ,重组融合型表达载体nCT/ pGEX 2T转化E coliBL2 1,培养表达 ,SDS PAGE分析可见Mr 约 310 0 0的融合蛋白带 ,融合蛋白占细胞内总蛋白的 34%。WesternBlotting分析证明该新型降钙素获得表达。
The novel hCT analogue was designed by Computer Aid Design Technology(CAD) The novel calcitonin gene(126bp)was synthesized by the combination of chemical synthesis and Klenow polymerase extension The synthetic gene was inserted into fusion expression vector pGEX 2T with BamHⅠ/EcoRⅠ sites This plasmid was transformed into E coli BL21 strain. The major band of about 31 000 fusion protein could be detected by SDS PAGE analysis and the fusion protein is more than 34% of total cellular protein The novel hCT analogue was confirmed by Western Blotting analysis
出处
《药物生物技术》
CAS
CSCD
1999年第3期129-134,共6页
Pharmaceutical Biotechnology
基金
国家科技部生命科学技术中心博士基金!(95 B 11)
关键词
降钙素
新型降钙素
基因工程
融合表达
Calcitonin ,Human calcitonin analogue,Genetic engineerring,Fusion expression
