人工合成抗菌肽D2A21基因的克隆和表达

为了获得高效表达的抗菌肽D2A21,本研究根据抗菌肽D2A21的氨基酸序列及大肠杆菌偏爱的密码子,设计并人工合成D2A21基因,并将该合成基因克隆至pProEXHTb载体中,构建成含有含6个组氨酸标签的重组质粒。重组质粒转化至大肠杆菌中,经摇瓶发酵,IPTG诱导后,发酵液用聚丙烯酰胺凝胶电泳分析。结果表明,成功合成D2A21基因,并构建了抗菌肽D2A21表达载体。通过镍柱亲和层析纯化,聚丙烯酰胺凝胶电泳分析,抗菌肽D2A21能够在大肠杆菌中稳定表达。本研究将为大规模表达纯化D2A21及其进一步的研究和利用提供一定基础。

Artificial synthesis,Cloning and Expression of Cecropin D2A21 Gene

To obtain stable multicopy cecropin D2A21,based on the established amino acid sequence of D2A21 and preferred codons of Escherichia coli,D2A21 gene sequence was designed and artificial synthesized.It was cloned into the integrative plasmid pProEXHTb to construct a recombinant expression vector containing histidine tag.The recombinant expression plasmid was transformed into E.coli10B,and the transformants were selected on the selection agar.After fermenting and incubating with IPTG,the fermentation product displayed by SDS-PAGE.The results showed that cecropin D2A21 gene was synthesized and cloned into expression plasmid successfully.Using Ni-NTA column and SDS-PAGE analysis clearly indicates that D2A21 was expressed in E.coli 10B successfully,then it would provides a basis for large scale expression and purification of D2A21,and also for its further research and utilization.