一种基于琼脂糖凝胶电泳法检测SARS-CoV-2的新方法  

A New Method for Detecting SARS-CoV-2 Based on Agarose Gel Electrophoresis

作　　者：刘军花 李学龙 刘茜阳 于琳 吴珊珊 尹秀山 LIU Jun-hua;LI Xue-long;LIU Qian-yang;YU Lin;WU Shan-shan;YIN Xiu-shan(Schl.of Ptuirm.&Bio-Engin.,Shenyang Uni.of Chem.Technol.,Shenyang 110142;BIOTECH(Shenyang)Biomed.Group Co.,Ltd.,Shenyang 110142;Jiangsu Plumway Med.Technol.Co.,Ltd.,Xuzhou 221000;Shanghai Nano Biotech.Co.,Ltd.,Shanghai 200000;BIOTECH(Jiangxi)Biotech.Co.,Ud.,Ganzhou 341000)

机构地区：[1]沈阳化工大学制药与生物工程学院,辽宁沈阳110142 [2]拜澳泰克(沈阳)生物医学集团有限公司,辽宁沈阳110142 [3]江苏谱录瑞医疗科技有限责任公司,江苏徐州221000 [4]海耐诺生物科技有限公司,上海200000 [5]拜澳泰克(江西)生物科技有限公司,江西赣州341000

出　　处：《微生物学杂志》2020年第3期51-54,共4页Journal of Microbiology

基　　金：2020年辽宁省重点研发计划项目(1580441949000);赣州市第一批新型冠状病毒感染的肺炎疫情应急科技攻关计划项目。

摘　　要：鉴于琼脂糖凝胶电泳分析方法的准确、快速、简便等特性,优化后可以做为检出COVID-19病原的新方法。在不影响结果判读的情况下,通过对扩增时间及检测样本浓度进行优化,以达到预期目的。结果表明,检测SARS-CoV-2的最优反应程序为94℃预变性3 min;94℃变性时间、56℃退火时间及72℃延伸时间均降至5 s,且进行35个循环;最后72℃孵育5 min,最优模板用量比例(初始值设为4 ng)下限调整在(1:1000)^(1:5000)范围内。阳性样本测试结果符合预期。建立了一种简便、省时,适用于检测低载量病毒样本的新方法,为临床提供诊断参考。In view of high accuracy,rapidity and simplicity of the electrophoretic analysis method,an optimized method for detecting of COVID-19 virus was provided.Without affecting the interpretation of the results,the amplification time and the concentration of the samples were optimized to achieve the expected purpose.The results showed that the optimal reaction procedure for detecting SARS-CoV-2 was firstly,predenaturation at 94℃for 3 min;then,the denaturation time at 94℃,annealing time at 56℃and elongation time at 72℃were all reduced by 5 s,and repeating for 35 cycles;finally,incubating at 72℃for 5 min will do.And the lower limit of the optimal reverse template consumption ratio(at initial value of 4 ng)can be adjusted to within 1/1000-1/5000.At the same time,the results of the positive sample test were tallied with expectations.Therefore,this study had established a new method with simple,time-saving and suitable for the detection of virus samples and with low-loading to provide diagnostic basis for further clinic.

关 键 词：COVID-19(新型冠状病毒感染的肺炎-2019) SARS-CoV-2(严重急性呼吸系统综合症冠状病毒2) PCR ORF1a/b基因 同源性 琼脂糖凝胶电泳法 

分 类 号：Q93-31[生物学—微生物学]