猪IFN-β启动子及其NF-κB结合位点萤光素酶报告载体的构建与活性检测

Construction and identification of luciferase reporter gene vectors directed by porcine IFN-β promoter and its NF-κB binding site

通过PCR的方法从猪基因组DNA中克隆IFN-β基因启动子片段,分别构建了含有猪β干扰素基因启动子萤光素酶报告质粒及其4个重复的NF-κB结合位点序列的萤光素酶报告质粒。脂质体基因转染法将萤光素酶报告质粒转染PK-15细胞,在poly(I∶C)或poly(dAT∶dAT)的刺激下,萤光素酶表达显著增加。本试验为进一步探讨猪β干扰素信号转导通路的研究奠定了基础。

To establish a method for detection of porcine proteins and genes related to IFN-β signal transduction, we analyzed the regulatory elements that regulate the transcription of porcine IFN-β gene. The promoter region of porcine IFN-β gene and its four copies NF-κB binding site regions were amplified from porcine genomic DNA by PCR and were cloned into promoter-free plasmid pGL3 basic. Then these constructs were transiently transfected into PK- 15 cells and luciferase activities were measured with or without the transfection of poly（I ： C） or poly（dAT ： dAT）. Higher expression of luciferase was obviously detected in PK-15 ceils transfected with poly（I ： C） or poly（dAT ： dAT）. These reporter constructs are important tools for investigation of porcine IFN-κB signaling transduction pathways.