optimization of cloning conditions for high-level production of recombinant mouse interleukin-2 in escherichia coli

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.