Optimization of Cloning Conditions for high-level Production of Recombinant Mouse Interleukin-2 in Escherichia coli

Backgrounds and objectives: Interleukin 2 (IL-2) secreted by activated CD4+ T cells  has been known as a major mediator in both  adaptive and native immune system  due to a board range of effects on  different cells in the immunity system (1-6). Methods: cDNA synthesis was performed using gene- specific primers designed by Gene Runner software 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 competent TOP10 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. 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 has produced in BL21 (DE3) pET-21b (+) expression system at 24°C in the soluble form.