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- Title
重组牛IFN-ω12生物学特性及其免疫调节机制的研究.
- Authors
安 东; 田其真; 曹 斌; 刘 静; 周 宁; 吉大庆; 卢 炜
- Abstract
A novel bovine IFN-ω12 was cloned and expressed, the protein stability and biological activity were analyzed in this study. Firstly, bovine IFN-ω12 gene was cloned from bovine liver genome and clonal vector was constructed. Bovine IFN-ω12 mature peptide encoding gene was amplified from clonal vector, and the prokaryotic expression vector pET-32aBoIFNω12 was constructed, then prokaryotic expression vector pET-32a-BoIFNω12 was constructed, and the recombinant vector was transformed into to express the fusion protein rBoIFNω12. The biological activity of the purified protein was detected in different cell lines. Results showed that rBoIFNω12 had antiviral activity against VSV on MDBK cells and PK-15 cells. In addition, rBoIFNω12 at 40μg/mL was less cytotoxic on MDBK cells than rBoIFNαA. After acid-base treatment or high temperature treatment, rBoIFNω12 still showed antiviral activity. The effect of rBoIFNω12 protein on key elements of interferon signaling pathway was detected by dual-luciferase assay on BL cells. The results showed that rBoIFNω12 could effectively activate the luciferase activity regulated by NF-ĸB response element, ISRE combining element and BoIFN-β promoter. Fluorescence guantitative PCR and western blot assay were used to detect the expression of ISGs in MDBK cells treated with rBoIFNω12, the results showed that the mRNA transcription and expression level of ISG15, ISG56 and Mx-1 were gradually increasing over time. The expression of Mx-1 was significantly higher than that of the untreated group. This study revealed that BoIFN-ω12 has antiviral and anti-proliferating activities with low cytotoxicity. It can stimulate the production of downstream proteins Mx1, ISG15, ISG56, and activate the luciferase activity activated by the promoter of NF-ĸB, ISRE, and BoIFN-β, has the typical characteristics of type I interferon, which can not only be a potential candidate for a novel, effective therapeutic agent, but also facilitate further research on the role of bovine IFN system.
- Publication
Chinese Journal of Preventive Veterinary Medicine / Zhongguo Yufang Shouyi Xuebao, 2023, Vol 45, Issue 7, p737
- ISSN
1008-0589
- Publication type
Academic Journal
- DOI
10.3969/j.issn.1008-0589.202209029