| ,2014,菜蛾斑蝥素受体PP2A催化亚基基因的原核表达与纯化[J].环境昆虫学报,(5):711-717 |
| 菜蛾斑蝥素受体PP2A催化亚基基因的原核表达与纯化 |
| Prokaryotic expression and purification of cantharidin receptor protein，phosphatase 2A catalytic subunit，from Plutella xylostella |
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| DOI： |
| 中文关键词: 小菜蛾 PP2Ac 原核表达 蛋白纯化 |
| 英文关键词:Plutella xylostella(L.) PP2Ac prokaryotic expression purification |
| 基金项目:国家自然科学基金项目 (30971938，31272099)；北京市高校人才强教计划项目 (PHR-201107135)；国家科技支撑计划课题 ( 2012BAD19B06) |
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| 中文摘要: |
| 为获得菜蛾斑蝥素受体PP2Ac的蛋白，支持斑蝥素受体结合机理等相关研究，本试验以菜蛾cDNA为模板，利用PCR方法扩增得到菜蛾PP2Ac基因，将PP2Ac基因连接至经Hind III与Bam HI双酶切处理的pET-30a原核表达栽体中，获得重组质粒并将其命名为pET30a-PX-PP2Ac。将pET30a-PX-PP2Ac化至大肠杆茵BL21中进行诱导表达获得的目的蛋白后，使用Ni-琼脂糖柱纯化对重组蛋白进行纯化。结果表明，菜蛾PP2Ac基因可以在大肠杆菌中表达，目的蛋白在不同的温度下均以包涵体形式存在。优化后的诱导表达条件是IPTG浓度0.2mM和温度30℃，目的蛋白大小约38KD。 |
| 英文摘要: |
| In order to obtain cantharidin receptor protein of Plutella xylostella (L.), in this experiment, PP2Ac gene were amplified by utilizing the DNA as template from the Plutella xylostella(L.), The PP2Ac gene was inserted to the prokaryotic expression vector pET-30a which was double enzyme restricted by Hind III and Bam HI. The recombinant vector，pET30a-PX-PP2Ac，was transformed into BL21 to induce PP2Ac protein expression. The target protein was purified according to Ni-Agarose Resin. The results showed, Plutella xylostella(L.) PP2Ac gene can be expressed in E. coli, but the target protein exist in the form of inclusion bodies at different temperatures. The optimum induction conditions of pET30a-PX-PP2Ac were as follows：IPTG concentration of 0.2mmol／L, induction temperature of 30℃。The purified product was about 38 kD that same as experimental expected result. |
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