【摘要】：目的建立馬爾堡病毒的環(huán)介導等溫擴增(loop-mediated isothermal amplification,LAMP)可視化快速檢測方法。方法人工合成馬爾堡病毒保守的核蛋白(Nucleoprotein,NP)編碼基因作為陽性標準品,設計LAMP擴增引物,以HNB作為可視化指示劑,通過LAMP濁度儀測定方法的靈敏度和特異性。并與常規(guī)PCR、實時熒光定量PCR作比較。結果建立的LAMP方法的靈敏度為100拷貝/μl,高于常規(guī)PCR及實時熒光定量PCR的靈敏度10~3拷貝/μl和10~2拷貝/μl,方法的特異性好,僅馬爾堡病毒有擴增曲線且HNB指示劑顏色發(fā)生陽性改變,其他病毒擴增均為陰性。體系的擴增效率較高,試驗僅需30min。結論建立的HNB-LAMP檢測方法簡便、快速、靈敏、特異,適用于馬爾堡病毒的可視化快速檢測。
[Abstract]:Objective to establish a rapid and visualized method for the detection of Marburg virus by ring-mediated isothermal amplification (loop-mediated isothermal amplification,LAMP). Methods the conserved nucleoprotein (Nucleoprotein,NP) coding gene of Marburg virus was synthesized as positive standard. LAMP primers were designed and HNB was used as visual indicator. The sensitivity and specificity of the method were determined by LAMP turbidimetry. And compared with conventional PCR, real-time fluorescence quantitative PCR. Results the sensitivity of the established LAMP method was 100 copies / 渭 l, which was higher than that of conventional PCR and real-time fluorescence quantitative PCR. The sensitivity of the method was 103 copies / 渭 l and 102 copies / 渭 l respectively. The specificity of the method was good. Only Marburg virus had amplification curve and the color of HNB indicator had positive change. All other virus amplification was negative. The amplification efficiency of the system was high, and the experiment only needed 30 minutes. Conclusion the established HNB-LAMP detection method is simple, rapid, sensitive, specific and suitable for rapid visual detection of Marburg virus.
【作者單位】： 中國藥科大學生命科學與技術學院;南京軍區(qū)軍事醫(yī)學研究所;
【基金】：國家重大傳染病防治專項(No.2013ZX10004103,2013ZX10004218) 江蘇省科技支撐計劃(社會發(fā)展)項目(No.BE2013603) 軍隊重點項目(No.BWS14J025)
【分類號】：R373;R440

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