重组Taq DNA聚合酶的表达和纯化鉴定

【目的】表达、纯化Taq DNA聚合酶,并检测其扩增性能。【方法】活化含Taq DNA聚合酶基因的菌株JM109,IPTG诱导表达。目的蛋白利用AKTA蛋白纯化系统,依次过Affi-Gel Blue Sepharose,Heparin Sepharose Fast Flow,Q-Sepha-rose Fast Flow,经SDS-PAGE分析,以国外进口Taq酶为标准,采用对比法初略测定酶活性,验证产品质量。【结果】制备的Taq DNA聚合酶具有扩增效率高、纯度高、特异性强、无核酸酶污染、活性稳定等优点。【结论】成功表达纯化Taq DNA聚合酶,其扩增性能达到甚至超过国外同类产品。

Expression, Purification and Identification of Recombinant Taq DNA Polymerase

In order to express and purify Taq DNA polymerase,and detect its amplification performance,E.coli strain JM109 containing the gene of Taq DNA polymerase was activated,and the gene was expressed under the inducement of IPTG.The expressed protein was purified by AKTA protein purification system through passing Affi-Gel Blue Sepharose,Heparin Sepharose Fast Flow and Q-Sepharose Fast Flow in turn.Taking Taq enzyme imported from abroad as the standard,the enzyme activity was roughly determined through antitheses and SDS-PAGE analysis,and the product quality was verified finally.The results revealed that the prepared Taq DNA polymerase had the following good characteristics: high amplification efficiency,high purity,strong specificity,no nuclease pollution and stable activity.In conclusion,the recombinant Taq DNA polymerase was successfully expressed and purified,and its amplification performance could reach or exceed that of the similar products imported from abroad.