Cloning of Streptococcus pyogenes streptokinase gene in E.coli
Subject Areas : Microbiologyelham siasi-torbati 1 , fatemeh fakhrefatemi 2 , میترا صاحی 3
1 - Faculty of Biological Sciences, Tehran-North Branch, Islamic Azad University, Tehran, Iran
2 - Department of Biotechnology, Collage of science, North Tehran Branch, Islamic Azad University, Tehran, Iran
3 -
Keywords: Cloning, E.coli XL1Blue, ska gene, Streptococcus pyogenes, Streptokinase.,
Abstract :
Aim and Background: Streptokinase is a thrombolytic medication and enzyme and is used to break down clots in some cases of myocardial infarction (heart attack), pulmonary embolism, and arterial thromboembolism. Recently recombinant streptokinase is important for treatment. In this research, was studied cloning of Streptococcus pyogenes streptokinase gene in E.coli. Materials and methods: At the first, genome of the bacteria was extracted and used PCR for ska gene. Then agarose gel electrophoresis was performed for detection of size and quality of the resulting amplicons. Then by TA Cloning kit, the target gene was inserted in to the PTG19 Cloning vector and the recombinant plasmid was inserted in to the E.coli XL1Blue bacteria. Checking the sequencing data obtained from cloned gene in the vectore compared with the refrenece sequenece obtained from the NCBI database, was confirmed the success of the gene cloning. Results: The results showed that PCR product is ska gene with length 513 bp. This is for the first time that the ska gene was cloned by the TA Cloning in the PTG19 vector and the E.coli XL1Blue host cell. The results of sequencing was confirmed presence of Streptokinase gene, ska. Conclusion: The ska gene was amplified and its correct and effective cloning was confirmed by TA Cloning with PTG19 vector and E.coli XL1Blue host cell.
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