miRNA“海绵”重组质粒的构建及对miR-483-5p的敲减  

MicroRNA sponges knockdown miR-483-5p in Hepa1-6 cells

作　　者：马宁[1] 王曦迪[1] 乔瑜[1] 李福源[1] 惠洋[1] 邹朝霞[1] 周凌云[1] 高旭[1] 

机构地区：[1]哈尔滨医科大学生物化学与分子生物学教研室,黑龙江哈尔滨150081

出　　处：《哈尔滨医科大学学报》2011年第1期1-4,共4页Journal of Harbin Medical University

基　　金：国家自然科学基金资助项目(81070457);黑龙江省自然科学基金青年基金资助项目(QC2010002)

摘　　要：目的建立microRNA失功能(loss of function)研究模型,为功能研究提供实验材料。方法针对miR-483-5p成熟体序列设计含6个拷贝反义序列的DNA片段,称为"S-483",将其连接入含CAG启动子的pCAGGs载体中,构建pCAGGs-S-483重组质粒。利用脂质体技术瞬时转染Hep1-6细胞,经re-al-time PCR对成熟miR-483-5p含量进行检测。结果转染pCAGGs-S-483组miRNA-483含量(0.33±0.04)较转染空质粒组(1.00)明显下降(P<0.01),但对其它miRNAs无影响。结论 S-483海绵策略可有效"吸附"并降低miR-483-5p,成功构建了pCAGGs-S-483重组质粒,为其功能研究和miRNA敲减动物的建立提供了有益借鉴。Objective To study microRNA function in vitro,a genetic strategies is required to generate loss-of-function phenotype.Methods The recombinant plasmids which expressed ＂microRNA Sponges＂,a six copies antisense sequence to mature microRNA miR-483-5p,were constructed.The miR-483-5p sponges plasmids were transient transfected into mouse Hepa1-6 cells.Real-time PCR was performed to identify the amounts of mature miR-483-5p.Results The amount of mature miR-483-5p was knockdowned（0.33±0.04） by the microRNA sponges recombinant plasmids with control group as 1.00（P0.01）.Conclusion The microRNA sponges strategies to knockdown microRNA is effective in loss-of-function research.

关 键 词：miRNA海绵 miR-483-5p 敲减 

分 类 号：Q78[生物学—分子生物学]