布鲁菌感染细胞NF-κB的表达水平检测及其干扰方法研究  

Comparative research of different interference techniques of NF-kappa B in Brucella infected cells

作　　者：荆明龙 李敏[1] 王小凤 李明奇[1] 霍凤娇 张豫[1] 刘良波[1] 张静[1] 张辉[1] Jing Minglong;Li Min;Wan Xiaofeng;Li Mingqi;Huo Fengjiao;Zhang Yu;Liu liangbo;Zhang Jing;Zhang Hui(College of Animal Science ＆ Technology,Shihezi University,Xinjiang,Shihezi 832000,China)

机构地区：[1]石河子大学动物科技学院,新疆石河子832000

出　　处：《石河子大学学报（自然科学版）》2018年第3期315-320,共6页Journal of Shihezi University(Natural Science)

基　　金：新疆兵团国际科技合作项目(GJHZ201709);新疆兵团优青计划(CZ027202);国家大学生SRP项目(201710759011)

摘　　要：目的本文针对布鲁氏菌侵染宿主细胞过程中NF-κB的表达进行检测,并比较siRNA和shRNA对RAW264.7细胞中NF-κB表达沉默效果。方法根据NF-κB P65基因序列,设计siRNA干扰序列,通过干扰RAW264.7细胞筛选效率较高的siRNA设计shRNA序列,siRNA和shRNA同时干扰布鲁氏菌侵染的RAW264.7细胞,使用RT-PCR方法检测NF-κB P65基因的相对表达情况。结果与阴性对照mock组相比,P65-shRNA组表达降低;P65-siRNA组P65表达低于P65-shRNA组,均低于阴性对照mock组;布鲁氏菌侵染组,P65表达显著高于阴性对照组。结论布鲁氏菌侵染组促进了NF-κB P65的表达,P65-shRNA、P65-siRNA组均抑制了NF-κB P65的表达,siRNA可作为基因沉默中快速筛选的工具,在RNA干扰的应用中可与shRNA配合使用。Objective The purpose of this research was to detect the expression of NF-κB P65 gene in Brucella infected host cells and compare the silencing effect of NF-κB expression by siRNA and shRNA interfered with RAW264.7 cells. Methods The siRNA interference sequences based on the NF-κB P65 gene sequence was designed. Through siRNA interfering with RAW264.7 cells, high interfering efficiency siRNA to design the shRNA sequence was screened. Both siRNA and shRNA interfered with Brucella infected RAW264.7 cells, NF-κB P65 gene relative expression was detected by RT-PCR. R esults Compared with the negative control mock group the expression of P65-shRNA decreased. The expression of P65 in the P65-siRNA group was lower than that in the P65-shRNA group, which was lower than that in the negative control mock group. Conclusion The expression of P65 in the Brucella infection group was significantly higher than that in the negative control group. The results showed that the Brucella infection group promoted the expression of NF-κB P65, P65-shRNA,P65-siRNA group all inhibited the expression of NF-κB P65, siRNA can be used as a rapid screening tool in gene silencing,in RNA interfering experiments shRNA could be coordinated.

关 键 词：布鲁氏菌 SIRNA SHRNA NF-ΚB 

分 类 号：S939.34[农业科学—渔业资源]