Aim and Background: Thermostable DNA polymerase enzyme has been taken much into consideration due to its application in PCR and molecular biology researches and it has doubled the importance of the study on the various thermostable DNA polymerase. The purpose of this study was to assay the function and efficient production of the cold sensitive thermostable DNA polymerase and its quick and cheap purification.
Material and Methods: After synthesis of the designed gene artificially, it was cloned into the pET28 vector and then was transferred into E.coli. IPTG was used as an inducer in the study of gene expression. Initial purification of the enzyme was performed by heat treatment at 72 °C and precipitation of other insoluble proteins by centrifuge. The DNA synthesis activity of crude extract was compared with commercial enzyme.
Result: Recombinant cold sensitive and thermostable Taq DNA polymerase expressed in E.coli showed a significant advantage and more desirable functionality such as activity and thermostable compared to commercial enzyme.
Conclusion: With regard to importance and application level of thermostable DNA polymerase in molecular biology, the production method used in this study is practical and cost effective. Additionally, the simplicity of producing method of this enzyme and its accuracy is a good reason for artificial and local production of highly pure cold sensitive Taq DNA polymerase.
