畜牧兽医学报2018，Vol.49Issue(6)：1282-1290,9.DOI:10.11843/j.issn.0366-6964.2018.06.021
猴源cathepsin基因shRNA文库及其稳定敲减细胞系的构建
Construction of shRNA Library of Chlorocebus Sabaeus Cathepsin Genes and the Establishment of Its Stable Transfection Cell Lines
摘要
Abstract
Up to now,the roles of cathepsins in the life cycle of Porcine reproductive and respirato-ry syndrome virus(PRRSV)remain unclear.This study was conducted to characterize the mech-anisms of cathepsins involved in PRRSV life cycle including adsorption,invasion,uncoating,bio-synthesis,assembly and release.The Chlorocebus sabaeus embryonic kidney epithelial cells Marc145(PRRSV can be propagated)derived from MA-104 was selected.Each cathepsin shRNA clone was constructed within the lentivirus plasmid vector pLKO.1,and the shRNA library con-sists of sequence verified shRNA lentiviral plasmid DNA against all the cathepsin genes that allow for high throughput loss-of-function screens.Puromycin was used to select inserts in mammalian cells.Finally,there were 45 clones involving 15 cathepsin genes in this cathepsin-shRNA library,and the expression of cathepsin mRNA in the stable cell lines was markedly downregulated.These stable cell lines will lay the foundation for the further study on the roles of cathepsins in PRRSV life cycle.关键词
cathepsin/shRNA/Marc145/慢病毒/稳定敲减细胞系Key words
cathepsin/shRNA/Marc145/lentivirus/stable knockdown cell line分类
农业科技引用本文复制引用
魏宇双,杨国宇,刘姣扬,韩莹倩,郭珍珍,刘晓贺,巴根,韩立强,王江,褚贝贝..猴源cathepsin基因shRNA文库及其稳定敲减细胞系的构建[J].畜牧兽医学报,2018,49(6):1282-1290,9.基金项目
国家自然科学基金(31490601 （）
31502031) （）
霍英东教育基金会青年教师基金项目(151033) （151033）
