DSN结合CHA用于miRNA的可视化检测  

DSN combined CHA for miRNA visual detection

作　　者：王奎宇 卜胜君 王月[1,2] 常洪标 孙秀伟 张红梅 万家余[2] WANG Kuiyu BU Shengjun WANG Yue CHANG Hongbiao SUN Xiuwei ZHANG Hongmei WAN Jiayu(Heilongjiang Bayi Agricultural University, Daqing 163319 ,China Institute of Military Veterinary, Academy of Military Medical Sciences, Changchun 130122,China Jilin Agricultural University, Changchun 130118 ,China)

机构地区：[1]黑龙江八一农垦大学,黑龙江大庆163319 [2]军事医学科学院军事兽医研究所,长春130122 [3]吉林农业大学,长春130118

出　　处：《黑龙江畜牧兽医》2017年第10期13-16,20,285,共6页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家重点研发计划项目(2016YFD0501001);吉林省自然科学基金项目(20150101105JC;20140101027JC)

摘　　要：为了研究灵敏特异、方便快捷的新型microRNA(miRNA)检测技术,试验采用双链特异性核酸酶(DSN)结合CHA(catalyzed hairpin assembly)恒温核酸扩增技术产生的双重信号放大方法,以试纸条作为信号输出方式用于miRNA的检测,将含有miRNA识别部分和CHA启动序列的复合探针与靶标miRNA结合后,DSN循环反应酶切释放CHA的启动序列T-trigger并引发CHA反应,最终产物可在试纸条检测线上留下红线。结果表明:本体系最低检测限可达到7.66 pmol/L,灵敏性好,且除靶标miRNA-21外,其余非特异性靶标均不能使试纸条留下红线,即使单碱基也能够被很好地区分,特异性强。说明本研究成功建立了等温、灵敏、特异的miRNA检测新方法,可用于miRNA的便携分析。The aim of the present study was to identify a new microRNA （miRNA） technology that was a highly sensitive, specific and conven- ient miRNA assay. A duplex specific nuclease （DSN） -catalyzed hairpin assembly （CHA） dual signal amplification method was applied in this experiment. The results of the miRNA detection were output through the test strip. After binding of the complex probe containing the miR- NA recognition sequence and the CHA promoter sequence to the target miRNA, the CHA reaction was initiated by the DSN cycling reaction to release the CHA promoter sequence T - trigger. The final products could leave a red line on the test strip. The results showed that the minimum detection concentration of miRNA was 7.66 pmol/L in the system. Except for the target miRNA - 21, the red line on the test strip could not be stayed by other non - specific targets. The single base could also be effectively recognized. Thus, a new method of isothermal, sensitive and specific miRNA detection was successfully established. The method could be conveniently used for miRNA analysis.

关 键 词：microRNA(miRNA) 双链特异性核酸酶(DSN) CHA 试纸条 可视化检测 

分 类 号：Q814[生物学—生物工程]