[Expression of recombinant extracellular region of human interleukin-1 receptor type I in Pichia pastoris].
OBJECTIVE
To construct pPICZalphaA-soluble interleukin-1 receptor type I (sIL-1RI) recombinant expression vector containing the gene fragment encoding the extracellular domain of sIL-1RI for its expression in Pichia pastoris.


METHODS
sIL-1RI gene was amplified by RT-PCR and inserted into the yeast expression vector pPICZalphaA by digestion ligation. The recombinant plasmid pPICZalphaA-sIL1RI was transformed into E.coli Stb13, and the positive clones were analyzed by PCR and DNA sequencing. The pPICZalphaA-sIL1RI recombinant plasmid was electroporated into GS115 cells and the transformants were analyzed by PCR. After phenotype identification, the recombinant strains were induced by methanol to express the target protein, which was analyzed by Western blotting of the cell extract and supernatant.


RESULTS
The recombinant plasmid pPICZalphaA-sIL-1RI was constructed successfully, and the results of Western blotting showed that yeast induced by methanol expressed a protein of about 39 kD.


CONCLUSIONS
sIL-1RI protein has been successfully expressed in P.pastoris expression system, which provides the basis for further study of sIL-1RI.
Lü
Wei-hui
WH
DME Center of Guangzhou Medical University of TCM, Guangzhou 510405, China. weihui.lu@gmail.com
Zhuang
Jun-hua
JH
Chen
Wei-ye
WY
Zhang
Zhan-feng
ZF
Huang
Xian-zhang
XZ
D004926:Escherichia coli / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D010843:Pichia / Q000378:metabolism*; D010957:Plasmids; D053573:Receptors, Interleukin-1 Type I / Q000096:biosynthesis* / Q000235:genetics*
C508590:IL1R1 protein, human; D053573:Receptors, Interleukin-1 Type I