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Published on: April 14, 2010
[Cloning and expression of human IL-1R II gene]
Yan-qiu Hu1, Jia-yin Liu, Chun Lu
1Department of Obstetrics and Gynecology, the First Affiliated Hospital, Nanjing 210029, China. huyanqiu78@163.com
Aim:
To clone human IL-1R II cDNA and construct its recombinant retrovirus vector so as to explore its role in IL-1R II related diseases.
Methods:
Human IL-1R II cDNA was amplified by RT-PCR from peripheral blood mononuclear cells (PBMCs) and inserted into the vector PET22b to construct recombinant vector PET22b-IL-R II. The recombinant was transfected into E. coli BL21 and expressed under IPTG induction. Expressed products were detected by Western blot. In addition, human IL-1R-II cDNA was subcloned into retrovirus vector LZRSPBMN and transfected into 293 cells by calcium phosphate precipitation. IL-1R II expression was detected by immunohistochemical staining.
Results:
IL-1R II cDNA with 1,203 bp was amplified by RT-PCR from human PBMCs. The recombinant of this cDNA could be expressed in E. coli,which was confirmed by Western blot results. Immunohistochemistry detection showed IL-1R II protein was expressed in 293 cells.
Conclusion:
Human IL-1R II gene was cloned successfully. PET22b-IL-1R II and LZR-IL-1R II were constructed and the recombinant protein IL-1R II was expressed in E.coli BL21. The results reported herein lay the foundation for further research on the role of IL-1R II in certain diseases.
