水产品和水体中灿烂弧菌现场可视化环介导等温扩增快检方法的建立及应用

Establishment and Application of Visual LAMP Methods for Quick Detection of Vibrio Splendidus in Aquatic Products and Water

基于环介导等温扩增(LAMP)技术建立水产品和养殖水域中灿烂弧菌现场可视化的快速、简便检测方法。以灿烂弧菌等作为研究对象,以灿烂弧菌的toxR基因作为靶基因,确定煮沸法为适合于弧菌基因组DNA提取的快捷方法,优化筛选的引物可以特异地检测灿烂弧菌,检测核酸浓度的灵敏度可以达到10^(-9) g/L,并且结果稳定、可靠。采用该方法进行实际样品的检测应用,在558份水体样品和655份水产品样品中,灿烂弧菌阳性检出率分别为0.54%和0.46%,与实时荧光PCR方法的检测结果符合率为100%。低技术要求,低设备成本以及短检测时间使得可视化LAMP快检方法成为现场可使用的一个理想选择,这对于水产养殖业来说也更方便。

A visualized,rapid,simple method was developed based on loop-mediated isothermal amplification(LAMP)to detect Vibrio splendidus in aquatic products and aquaculture waters.Taking Vibrio splendidus as the research object,and the toxR gene of Vibrio splendidus as the target gene,the boiling method was determined as a fast method suitable for the genomic DNA extraction of Vibrio splendidus.The optimized and screened primers could specifically detect Vibrio splendidus,and the sensitivity of nucleic acid concentration detection could reach 10^(-9)g/L.We show that the results were stable and reliable.In 558 water samples and 655 aquatic product samples,the positive detection rate of Vibrio brilliant was 0.54%and 0.46%,respectively,and the coincidence rate with the real-time PCR method was 100%.Low technical requirements,low equipment costs,and short detection time render the visual LAMP rapid detection method an ideal choice in field diagnosis and convenience for the aquaculture industry.