| چکیده انگلیسی مقاله |
Backgrounds Many proteins have been expressed so far in bacterial host. Due to its simple culture conditions, having a short life cycle, and easily genetic manipulation, E.coli have been regarded as a preferable host to produce recombinant proteins, but protein cloning in bacterial host have many challenges. Therefore, we aimed to review some of these problems by an experience from mice IL-2 recombinant. Materials and methods: cDNA synthesis was performed after RNA extraction of mouse splenocytes. PCR product purification carried out after IL-2 coding sequence amplification and was ligated into the pET-21b (+) vector and transformed into the competent BL21 E.coli. Expression and purification of recombinant mouse IL-2 were done using IPTG inducer and metal affinity chromatography respectively. Results: DNA sequencing confirmed the accuracy of the insertion process. A 23 kDa exogenous protein was observed on the SDS-PAGE. Specificity and concentration of produced mouse recombinant IL-2 protein were confirmed by western blotting and BCA methods. Conclusion: Recombinant IL-2 was produced in BL21 and pET-21b (+) expression system at 24°C in the soluble form. |
| نویسندگان مقاله |
| Arezou Abdi
ImmunoBiochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran
| Mitra Hosseinpour
ImmunoBiochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran
| Kazem Mashayekhi
ImmunoBiochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran
| Mohammad Javad Mousavi
Immunology Department, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran
| Seyedeh Elham Badiee Kheirabadi
ImmunoBiochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran
| Mojtaba Sankian
ImmunoBiochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran
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