Expression of Human Interleukin-33 and studies on its biology function
- VernacularTitle:人白细胞介素33基因的克隆表达及其生物学活性的研究
- Author:
Yanfang JIANG
;
Zhaoxia WANG
;
Guijie XIN
;
Yubin LI
;
Junqi NIU
- Publication Type:Journal Article
- Keywords:
IL-33;
Gene cloning and expression;
Bacillus coli;
Bioactivity
- From:
Chinese Journal of Immunology
1985;0(02):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To clone human interleukin-33(hIL-33)and express it in E.coli efficiently.Methods:The primers were synthesized according to the hlL-33 cDNA sequence in GeneBank.The hIL-33 was amplified by RT-PCR from human fibroblast cell line(L929),the PCR product was inserted into pUC19 vector.The IL-33 cDNA confirmed by sequencing was inserted into expressing vector PQE30 and expressed in E.coli M15 strain.IL-33 protein expression was induced by IPTG and purified by Ni-NTA affinity chromatography.The recombinant IL-33 was identified by Immunoblot and its biological activity was analyzed.Results:DNA sequencing confirmed that the cloned cDNA was identical to the published sequence of hIL-33.The recombinant plasmid PQE/hIL-33 was transformed into M15.An expected 18KD protein of hIL-33 found mainly in the induced host strains about 25% of total bacteria lysis by SDS-PAGE and coomassie blue staining.The 18 KD protein could be recognized by anti-IL33 antibody in western blot.The recombinant protein was purified to more than 95% of total protein and induces the production of IL-4 and IL-5 in human peripheral blood mononuclear cells.Conclusion:We have successfully expressed hIL-33 protein in E.coli and the expressed product has IL-33 specific bioactivity.
