دوره 7، شماره 1 - ( 11-1397 )                   جلد 7 شماره 1 صفحات 16-25 | برگشت به فهرست نسخه ها

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Abdi A, Hosseinpour M, Mashayekhi K, Mousavi M J, Badiee Kheirabadi S E, Sankian M. Optimization of Cloning Conditions for High-Level Production of Recombinant Mouse Interleukin-2 in Escherichia coli. Res Mol Med (RMM). 2019; 7 (1) :16-25
URL: http://rmm.mazums.ac.ir/article-1-309-fa.html
Optimization of Cloning Conditions for High-Level Production of Recombinant Mouse Interleukin-2 in Escherichia coli. Research in Molecular Medicine. 1397; 7 (1) :16-25

URL: http://rmm.mazums.ac.ir/article-1-309-fa.html


چکیده:   (793 مشاهده)
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.
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نوع مطالعه: پژوهشي | موضوع مقاله: ايمونولوژي
دریافت: ۱۳۹۸/۳/۳ | پذیرش: ۱۳۹۸/۴/۱۶

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