【摘要】 根据甜蛋白莫奈林的氨基酸序列,选择酵母偏爱密码子,化学合成单链莫奈林(Single ChainMonellin,SCM)基因片段,并拼接成全基因.基因的DNA序列分析表明,合成的单链莫奈林基因与设计的一致.该基因插入于毕赤氏酵母整合型质粒pPIC9K中置于AOXI启动子的控制下,并通过电转化技术将重组质粒pPIC9K/SCM导入Pichiapastoris,在含有G418的YEPD平板上筛选整合型的多拷贝的转化子.转化子在BMMY培养基中经甲醇诱导表达,用SDS PAGE检测发酵液,表明SCM的分泌表达蛋白可达发酵液蛋白含量的90%.更多还原

【Abstract】 According to the amino acid sequence of the sweet protein monellin,a gene of singlechain monellin(SCM) was synthesized by the chemical synthesis method. The sequence analysis shows that the synthesized SCM was accordant with the expected design. The expression vector was then constructed with the integrated vector pPIC9K,and transformed to Pichia pastoris SMD1168 using electroporation. The transformants was screened with RDB and G418 resistance. The SDSPAGE analysis indicates that there is a clear band of SCM secretion protein and the content of has achieved the approximately 90% of the total proteins.更多还原