摘要
构建了针对1.8-kb mRNA基因簇潜在阅读框的RNA干扰质粒pP(1.8-kb-RNAi)。将该质粒与包含其上游双向启动子的质粒pP(pp38)-CAT和pP(1.8-kb)-CAT共转染鸡胚成纤维细胞(CEF)和MDV rMd5感染的CEF(rMd5-CEF),48 h后,通过测定转染细胞裂解液中氯霉素乙酰转移酶(CAT)的活性确定1.8-kb mRNA被干扰后对双向启动子活性的影响。结果显示,利用pP(1.8-kb-RNAi)质粒干扰1.8-kb mRNA,可以使其上游双向启动子两个方向的活性均显著下降(P<0.01),其中1.8-kb mRNA方向下跌29.5%,pp38方向下跌25.0%。本研究结果证明了1.8-kb mRNA对其上游双向启动子活性有增强作用。
Marek＇s disease virus （MDV） contains a bi-directional promoters located between pp38 gene and 1.8-kb mRNA in the long inverted repeat region of the viral genome. The involvement of 1.8-kb mRNA in up-regulating the activity of these promoters was analyzed by transient expression of chloramphenicol acetyltransferase （CAT） reporter gene. A plasmid pP（1.8-kb-RNAi） was constructed to interference with 1.8-kb mRNA. Two CAT reporter plasmids, pP（pp38）-CAT and pP（1.8-kb）-CAT, were transfected with pP（1.8-kb-RNAi） into chicken embryonic fibroblast （CEF） and CEF infected with rMdS,respectively. No CAT activity was detected in uninfected CEF as ex- pected. The activity of the bi-directional promoter was significantly decreased in both directions when interfered by the plasmid pP（1.8-kb-RNAi）. 29.5% and 25.0% activity were decreased in the direction of 1.8-kb mRNA family and pp38 direction, respectively. It shows that 1.8-kb mRNA plays an important role in regulating the transcriptional activity of the bi-directional promoter.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2009年第11期1369-1372,共4页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(30700596)
