中国兽医科学2025，Vol.55Issue(4)：427-434,8.DOI:10.16656/j.issn.1673-4696.2025.0060
基于遗传密码子扩展技术的口蹄疫病毒VP1蛋白位点特异性荧光标记
Site-specific fluorescence labeling of VP1 protein of foot-and-mouth disease virus based on genetic code expansion technology
摘要
Abstract
To introduce fluorescent probe into VP1 protein of foot-and-mouth disease virus(FMDV)in living cells so as to achieve the dynamic visualization of protein in vivo,the unnatural amino acid(UAA)incorporated site of VP1 protein was firstly screened.Incorporated UAA into specific sites of VP1 protein using genetic code expansion technology,and fluorescent molecules DIBO-Alexa Fluor-488 were introduced by biological orthogonal click chemistry,then the staining of VP1 protein in PK-15 cells was observed.The fluorescently labeled proteins were further verified by indirect immunofluo-rescence assay.The amino acid mutation sites with high UAA insertion efficiency were screened suc-cessfully,and the expression of VP1 protein could be controlled by whether UAA was added or not.Fluo-rescent molecular dye DIBO-Alexa Fluor-488 and UAA on VP1 protein can be used for live cell staining,and immunofluorescence staining was performed at the same time.The staining by the new method of fluo-rescence labeling of VP1 protein can overlay with the traditional method of immunofluorescence.In con-clusions,a new method was used to achieve site-specifically labeling of FMDV VP1 protein in living cells,which laid a foundation for the study of VP1 protein function and tracing the infection process of FMDV in cells.关键词
蛋白质标记/口蹄疫病毒/VP1蛋白/点击化学/非天然氨基酸Key words
protein labeling/foot-and-mouth disease virus/VP1 protein/click chemistry/unnatu-ral amino acid分类
畜牧业引用本文复制引用
郝荣增,卢炳州,杨洋,毛玉涵,王姿逸,赵陇和,李亚军,茹毅,郑海学..基于遗传密码子扩展技术的口蹄疫病毒VP1蛋白位点特异性荧光标记[J].中国兽医科学,2025,55(4):427-434,8.基金项目
国家自然科学基金项目(32372990) （32372990）
甘肃省自然科学基金项目(22JR5RA030) （22JR5RA030）
