The development of loop-mediated isothermal amplification combined with lateral flow dipstick for detection of Karlodinium veneficum

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• 作者：Hai-Long Huang ^{baiduhailong@sina.com} ; Peng Zhu ; ^{zhupeng@nbu.edu.cn} ; Cheng-Xu Zhou ^{zhouchengxu@nbu.edu.cn} ; Shan He ^{heshan@nbu.edu.cn} ; Xiao-Jun Yan ^{yanxiaojun@nbu.edu.cn}
• 关键词：LAMP ; loop mediated isothermal amplification ; LFD ; lateral flow dipstick ; ITS ; internal transcribed spacer ; PCR ; polymerase chain reaction ; KV ; karlodinium veneficum ; AGE ; agarose gel electrophoresis ; EB ; ethidium bromide ; PSU ; practical salinity units ; H7N9 ; avian influenza virus ; FITC ; fluorescein isothiocyanate ; FIP ; forward inner primer ; BIP ; backward inner primer ; NJ ; neighbour-joining ; POCT ; point-of-care testing
• 刊名：Harmful Algae
• 出版年：2017
• 出版时间：February 2017
• 年：2017
• 卷：62
• 期：Complete
• 页码：20-29
• 全文大小：1788 K
• 卷排序：62

文摘

The aim of this study was to develop a loop-mediated isothermal amplification (LAMP) combined with a chromatographic lateral flow dipstick (LFD) assay to rapidly and specifically detect the Karlodinium veneficum ITS gene. Four groups of LAMP primers were specially designed to target the K. veneficum ITS gene. The LAMP-LFD detection limit was 7.4 pg/μL (approximately 6.5 cells/mL) of K. veneficum genomic DNA and was 10 times more sensitive than standard PCR. The LAMP-LFD method exhibited high specificity and accurately identified K. veneficum algal isolates, but not other algal isolates. To test the assay’s accuracy, samples from positive results were further analyzed by sequencing and phylogenetic analysis, all of which were identified as K. veneficum. Over all, the LAMP-LFD assay established in this paper can be used as a reliable and simple method to detect the K. veneficum.