兔防御素NP2基因克隆及其毕赤酵母表达的研究  

Cloning and Expression of Rabbit Defensin NP2 Gene in Pichia Pastoris

作　　者：李再新[1] 李光辉[1] 李昌泽[1] 江怀进[1] 杨英[1] 王惠[1] 

机构地区：[1]四川理工学院生物工程系,自贡643000

出　　处：《畜牧兽医学报》2005年第9期869-872,共4页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：四川理工学院课题基金项目

摘　　要：根据兔防御素NP2基因序列和Pichia酵母密码偏爱性,设计兔防御素NP2基因序列片段,构建pGAPZαA酵母表达重组子,转化E.coliJM109,扩增重组子,经限制性内切酶BlnI线性化,转Pichia酵母。PCR扩增挑选阳性转化子,SDS-PAGE电泳分析兔防御素NP2酵母表达、Bradford法蛋白质定量和抗菌试验。结果表明:兔防御素NP2可在Pichia酵母中表达,且对大肠杆菌具有显著的抗菌作用,为兔防御素NP2深入研究和开发利用奠定了一定的物质和理论基础。A DNA fragment encoding rabbit defensin NP2 was cloned based on the published amino acid sequence and the biased codon usage of Pichia Pastoris. The modified rabbit defensin NP2 DNA fragment was inserted into Pichia Pastoris expression vector pGAPZα. Then the plasmid pGAPZαA- NP2 was transferred to E. coli JM109 and used for Pichia pastoris transformation. The recombinant linearized with restriction enzyme BlnI was transformed into special strain Pichia pastoris SMD1168. The positive clones were selected with PCR, and the recombinant rabbit defensin NP2 was detected with SDS-PGAE, and quantified with Bradford and then used for biological activity assay. The results show that the modified rabbit defensin NP2 can express in the Pichia pastoris, with the well ability to inhibit the growth of E. coli 5α. These findings have provided valuable tools for further studies on the function and medicinal application of NP2.

关 键 词：防御素NP2 毕赤酵母表达 抗菌 

分 类 号：S829.12[农业科学—畜牧学]