Stable expression of mouse IFN-λ2 in CHO cells and its biological activity analysis
10.3760/cma.j.issn.0254-5101.2010.02.002
- VernacularTitle:鼠IFN-λ2 CHO细胞系建立及生物学活性的研究
- Author:
Yulan YAN
;
Lixue YUAN
;
Yang LIU
;
Wenyan CAO
;
Xuefeng BU
;
Zhigao BU
;
Jinxu ZHENG
- Publication Type:Journal Article
- Keywords:
mIFN-λ2;
Gene clone;
Stably expressing;
Antiviral activity;
Mx1 protein
- From:
Chinese Journal of Microbiology and Immunology
2010;30(2):104-109
- CountryChina
- Language:Chinese
-
Abstract:
Objective To express mouse IFN-λ2 stably and study its biological activity. Methods Full-length of mIFN-λ2 cDNA was obtained by using RT-PCR from cells of mouse spleen stimulated by ve-sicular stomatitis virus(VSV) and then subcloned to eukaryotic expressing vector PCAGG-EGFP. The recom-binant was transfected into CHO cells. VSV * GFP-B16 system was used to measure the antivirus activity. The constructed cell line MDBK-Mxp-Luc was used to study the character of Mx1 protein induced by the mIFN-λ2. Results The recombinant pMD18-T-mIFN-λ2 was digested by two kinds of enzyme, Sac I and Xho I, to produce the fragment was of 582 bp, and of which the sequence analysis of sequence shows it was entirely consistent with the nucleotide sequences reported in GenBank. PCAGG-EGFP-mIFN-λ2 eukaryotic expressing vector was constructed successfully and expressed stably in CHO cells, and the mRNA of mIFN-λ2 was verified expressing in CHO-PCAGG-EGFP-mIFN-λ2 cell line by RT-PCR. The antivirus activity of in the supernatant secreted by the CHO-PCAGG-EGFP-mIFN-λ2 cell line was 10~4 AU/ml. The mIFN-λ2 pro-tein can could induce the expression of the antivirus protein Mx1, and the expression of Mx1 protein induced by mIFN-λ2 enhanced with time going, 9 to 12 hours achieved the peak, 24 hours vanished. Conclusion Gene cloning of mIFN-λ2 was successful. The eukaryotic expressing vector of mIFN-λ2 was constructed suc-cessfully and expressed stably in CHO cells, and its product has obvious antivirus activity in vitro. And the antivirus activity of the product was closely correlated with inducing expression of antivirus protein Mx1.
