畜牧与兽医2025，Vol.57Issue(8)：43-48,6.
猪源NUB1的基因克隆、表达与细胞定位研究
Gene cloning,expression and cell localization of porcine NUB1
摘要
Abstract
The aim of this study was to identify,express and localize the negative regulator of ubiquitin-like proteins 1(NUB1),and to further study the effects of NUB1 on porcine rotavirus(PoRV)replication,innate immunity and the neural precursor cell-expressed develop-mentally downregulated 8(NEDD8)pathway regulation.Primers with Kpn Ⅰ and Xho Ⅰ restriction sites were designed,and His tag was in-troduced into the C-terminal.Using the cDNA of IPEC-J2 cell as a template,the NUB1 gene was amplified and cloned into a pcDNA3.1(+)vector.The recombinant plasmid obtained was transfected into MARC145 cells by Lipofectamine™ 3000 liposome.The expression of NUB1 in MARC145 cells was detected by Western blot and indirect immunofluorescence,and the localization of NUB1 was determined by confocal microscopy.The results were as follows:The porcine NUB1 gene was successfully amplified,and the eukaryotic expression plasmid pcDNA3.1(+)-NUB1-His was constructed.The recombinant plasmid could be expressed in MARC145 cells,and NUB1 protein was loca-ted in the nucleus and cytoplasm.To sum up,the eukaryotic expression plasmid pcDNA3.1(+)-NUB1-His,which can express NUB1 ex-ogenously,was successfully constructed in this experiment,which laid a foundation for further research on the biological function of NUB1 and its influence on PoRV replication.关键词
NUB1/NEDD8/基因克隆/细胞定位/基因表达Key words
NUB1/NEDD8/gene cloning/subcellular localization/gene expression分类
农业科技引用本文复制引用
卢洪婷,朱雪蛟,周金柱,张雪寒,陶然,郭容利,牛家强,李彬..猪源NUB1的基因克隆、表达与细胞定位研究[J].畜牧与兽医,2025,57(8):43-48,6.基金项目
国家重点研发计划项目(2023YFD1801303) （2023YFD1801303）
国家自然科学基金项目(32302892) （32302892）
江苏省自然科学基金项目(BK20221432) （BK20221432）
