Rapid and sensitive detection of white spot syndrome virus by loop-mediated isothermal amplification combined with a lateral flow dipstick

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• 作者：Wansadaj Jaroenram ; Wansika Kiatpathomchai ; Timothy W. Flegel
• 关键词：WSSV ; White spot syndrome virus ; Penaeus monodon ; Shrimp ; Loop-mediated isothermal amplification ; LAMP ; Nested PCR ; Lateral flow dipstick ; LFD
• 刊名：Molecular and Cellular Probes
• 出版年：2009
• 出版时间：April 2009
• 年：2009
• 卷：23
• 期：2
• 页码：65-70
• 全文大小：465 K

文摘

Loop-mediated isothermal amplification (LAMP) allows rapid amplification of nucleic acids under isothermal conditions using a set of four specifically designed primers that recognize six distinct target sequences. It can be combined with a chromatographic lateral flow dipstick (LFD) for highly specific, rapid and simple visual detection of WSSV-specific amplicons. Using this protocol, a 30-min amplification followed by 5 min hybridization with an FITC-labeled DNA probe and 5 min LFD resulted in visualization of DNA amplicons trapped at the LFD test line. Thus, 10 min for rapid DNA extraction followed by LAMP combined with LFD detection resulted in a total assay time of approximately 50 min. Detection sensitivity was comparable to other commonly-used methods for nested PCR detection of WSSV but had the additional advantages of reduced assay time, confirmation of amplicon identity by hybridization and elimination of electrophoresis with carcinogenic ethidium bromide.