A simplified protocol for producing Taq DNA polymerase in biology laboratory
| Autor: | Touraj Farazmandfar, Alireza Rafiei, Mohammad Bagher Hashemi-Sotehoh, Reza Valadan, Mohammad Alavi, Fatemeh Moradian |
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| Jazyk: | angličtina |
| Rok vydání: | 2013 |
| Předmět: | |
| Zdroj: | Research in Molecular Medicine, Vol 1, Iss 2, Pp 23-26 (2013) |
| Druh dokumentu: | article |
| ISSN: | 2322-1348 2322-133X |
| Popis: | Background: Taq DNA polymerase is a very important enzyme for molecular biological studies such as DNA amplification and DNA sequencing by the PCR. It is a standard enzyme that is used in 90% of molecular biology labs today. The aim of this study was to produce Taq DNA polymerase enzyme in E. coli by a reliable, practical, simple and low cost method.Materials and Methods: In this study, the Taq gene was amplified from the genomic DNA of Thermus aquaticus and cloned into pTrc99A vector. Recombinant plasmid is expressed in E. coli strain TOP10. Product protein is extracted and purified. Expression of gene was analyzed by SDS-PAGE and gene amplification.Results: SDS-PAGE showed an approximately 94 KDa protein. The density of protein bands in agarose gel electrophoresis indicated that the purified enzyme is more active than the non purified one.Conclusion: The protocols described in this paper lead to the production of pure and active enzyme that can be applied in both teaching and research laboratories. |
| Databáze: | Directory of Open Access Journals |
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