Article

Published date: Sep 30 2019

Journal Title:

Issue title: Research in Molecular Medicine (RMM): Volume 7, Issue No. 1

DOI: 10.18502/rmm.v7i1.5256

Arezou AbdiImmuno-Biochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran

Mitra HosseinpourImmuno-Biochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran

Kazem MashayekhiImmuno-Biochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran

Mohammad Javad MousaviDepartment of Hematology, Faculty of Allied Medicine, Bushehr University of Medical Sciences, Bushehr, Iran; Immunology Department, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran

Seyedeh Elham Badiee KheirabadiImmuno-Biochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran; Immunology, Torbat Heydariyeh University of Medical Sciences, Torbat Heydariyeh, Iran

Mojtaba Sankiansankianm@mums.ac.irImmuno-Biochemistry lab, Immunology Research Center, Mashhad University of medical Sciences, Mashhad, Iran

Backgrounds and objectives: 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. 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.