基于RT-RPA的微量核酸释放剂的研制  

Development of micro-nucleic acid releasing reagent based on RT-RPA detection method

作　　者：常宇桐 刘丽娟[1] 张丽萍[1] 胡孔新[1] CHANG Yu-tong;LIU Li-juan;ZHANG Li-ping;HU Kong-xin(Institute of Health Inspection and Quarantine,Chinese Academy of Inspection and Quarantine,Beijing 100176,China)

机构地区：[1]中国检验检疫科学研究院卫检所,北京100176

出　　处：《中国国境卫生检疫杂志》2022年第3期172-174,共3页Chinese Journal of Frontier Health and Quarantine

基　　金：国家市场监管总局技术保障专项(2020YJ046);中国检科院基本科研业务费项目(2020JK022)。

摘　　要：目的配制一种用于反转录重组酶聚合酶扩增(RT-RPA)方法的微量核酸释放剂,用于病毒的核酸检测。方法用异硫氰酸胍、Tween-20、TritonX-100、乙醇、异戊醇和无酶无菌水等配制成微量核酸释放剂,对几种成分浓度、配比进行优化,用诺如病毒GII型阳性粪便样本,结合RT-RPA和商品化试剂盒对微量核酸释放剂的效果进行验证。结果优化后的微量核酸释放剂组成及配比是:40%(50 mmol/L)异硫氰酸胍、0.05%Tween-20、0.05%TritonX-100、25%乙醇、20%异戊醇和14.9%无酶无菌水,和待检样本1∶1混合后用于RT-RPA检测,检测结果与商品化荧光定量试剂盒的结果一致,均能检测出诺如病毒GII型阳性样本。结论配制的微量核酸释放剂可用于病毒的核酸检测。Objective To prepare a micro-nucleic acid releasing reagent applied in RT-PRA method,to detect virus nucleic acid.Methods The micro-nucleic acid releasing reagent was composed of guanidine isothiocyanate,Tween-20,TritonX-100,ethanol,isoamyl alcohol and enzyme-free sterile water.The main components,their concentration and ratio were all optimized.The micro-nucleic acid releasing reagent was verified by using norovirus GII positive stool samples,combined with RT-RPA and commercial Real-time PCR kit.Results After optimized,the components of micro-nucleic acid releasing reagent was 40%(50 mmol/L)guanidinium isothiocyanate,0.05%Tween-20,0.05%Triton X-100,25%ethanol,20%isoamyl alcohol and 14.9%enzyme-free sterile water,mixed with samples with 1∶1 ratio for RT-PRA detection.The detection result was consistent with commercial Real-time PCR kit.Conclusion This micro-nucleic acid releasing agent can be used for virus nucleic acid detection.

关 键 词：核酸释放剂 重组酶聚合酶扩增 诺如病毒 

分 类 号：R183[医药卫生—流行病学] R373[医药卫生—公共卫生与预防医学]