本文關鍵詞：埃博拉病毒萊斯頓亞型實時熒光定量PCR檢測方法的建立　出處：《病毒學報》2015年03期 　論文類型：期刊論文

  更多相關文章： 埃博拉病毒萊斯頓亞型 熒光定量PCR 病毒載量

【摘要】：本研究的目的為建立一種靈敏、快速、定量檢測埃博拉病毒萊斯頓亞型核酸的實時熒光定量PCR檢測方法及試劑盒。首先選擇埃博拉病毒萊斯頓亞型的保守基因NP基因作為檢測靶目標,篩選出保守序列并設計合成一對特異性引物。然后將連有NP全長基因的重組質粒作為定量標準品,將10倍系列稀釋的重組質粒進行熒光定量PCR擴增,繪制標準曲線,并進行重復性、靈敏度及特異性檢測。結果顯示建立的埃博拉病毒萊斯頓亞型熒光定量PCR檢測方法,其靈敏度可達102拷貝/μL,不同梯度標準品間線性關系(R2)達0.997,斜率為-0.3101,擴增效率為110.145%,所有標準品均在79.94℃出現(xiàn)尖且窄的特異性熔解峰。利用該檢測系統(tǒng)可以快速定量檢測埃博拉病毒萊斯頓亞型核酸,靈敏度高、重復性好,可用于基礎及臨床實驗室對埃博拉病毒萊斯頓亞型感染的快速診斷和臨床效果的監(jiān)測,具有實際的應用價值。
[Abstract]:The aim of this study was to establish a sensitive and rapid method. Quantitative detection of Ebola virus Leston subtype nucleic acid real-time fluorescence quantitative PCR detection method and kit. First select the conserved gene NP gene of Ebola virus Leston subtype as the target detection target. The conservative sequence was selected and a pair of specific primers were designed and synthesized. Then the recombinant plasmid with NP full-length gene was used as the quantitative standard, and the 10-fold diluted recombinant plasmid was amplified by fluorescence quantitative PCR. The standard curve was drawn, and the repeatability, sensitivity and specificity were detected. The results showed that the sensitivity of the established fluorescent quantitative PCR method for detection of Ebola virus Leston subtype could reach 102 copies / 渭 L. The linear relationship between different gradients was 0.997, the slope was -0.3101, and the amplification efficiency was 110.145%. All the standard samples showed sharp and narrow specific melting peak at 79.94 鈩，

本文編號：1384955