禽源与鼠源U6启动子对鸡马立克氏病病毒 gIgE基因shRNA干扰活性的比较 毕业论文.doc

毕业论文 禽源与鼠源U6启动子对鸡马立克氏病Comparison of interfering effect of shRNA against Marek’s disease gI/gE gene under chicken or mouse U6 promoter 学 生： 指导教师： 院系： 动物科技学院 专 业： 班 级： 20级 论文提交日期：20年06月日目 录 引言 3 1 材料 4 1.1工程菌、细胞、质粒及毒株 4 1.2主要试剂和仪器 4 2 方法 5 2.1不同来源的U6启动子驱动siRNAs的效果评价 5 3 结果 5 3.1不同的启动子驱动siRNAs的效果评价 5 4 讨论 8 4.1禽源U6和鼠源U6在不同细胞中表达siRNAs的效率 8 5 结论 9 参考文献 10 致 谢 12 禽源与鼠源U6启动子对鸡马立克氏病）[1]引起的一种免疫抑制性肿瘤疾病。用已构建好的带有禽源启动子的重组质粒pcU6-shgI735和pcU6-shgE936和带有鼠源启动子的重组质粒pmU6-shgI735、pmU6-shgE936，随报告质粒pEGFP-gI或pEGFP-gE分别共转染DF1、MDCK、Vero细胞[2]，比较了禽源U6和鼠源U6启动子的驱动siRNAs[3]的转录活性。结果显示，在DF-1细胞上禽源U6驱动特异性siRNA对gI、gE[4]基因抑制率分别为87.39%和 85.04% ，其活性是哺乳动物细胞1.21-1.45倍，差异显著（p0.05））：gI基因；gE基因；siRNA Comparison of interfering effect of shRNA against Marek’s disease gI/gE gene under chicken or mouse U6 promoter Abstract:Mareks disease is an immune suppression tumosis caused by Marek s disease virus.The recombinant plasmids pcU6-shgI735, pcU6-shgE936 containing chicken U6 promoter and pmU6-shgI735, pmU6-shgE936 containing mouse U6 promoter were constructed previouly. Then these recombinant plasmids were co-transfected respectively into DF-1, Vero and MDCK cell line with plasmids pEGFP-gI or pEGFP-gE reporter plasmid respectively to measure gI and gE gene knock down efficiency on different cell lines and compare the shRNA expression efficiency driven by chicken U6 and mouse U6 promoter.This paper revealed that the knock down efficiency of pcU6-shgI735 plasmids on gI gene was 87.39%、76.19%、60.09% on DF-1, MDCK and Vero cells, respectively; while that of pcU6-shgE735 plasmids on gE gene was 85.04%、73.60%、58.57% respectively. The recombinant plasmid with the shRNA under the control of cU6 promoter was 1.21 to 1.45 times more efficient in CEF than that in mammalian cells (i.e. Vero or MDCK cells) for reducing the target gene expression.while the mouse U6 promoter transcription activity in mammalian cells was 1.24 to 1.54 times more higher than that