一种基于SYBR Green的实时荧光定量PCR检测方法用于检测环境水样中的DNA的开发。
Development of a SYBR Green-Based Real-Time PCR Assay to Detect DNA in Environmental Water Samples.
作者信息
Manalo Daria L, Bolivar Jude Karlo G, Ermino Karl Ian T, Bondoc Jeromir G, Espino Mark Joseph M, Panganiban Efraim P, Nogrado Kathyleen S, Fornillos Raffy Jay C, Jiz Mario A, Leonardo Lydia R, Fontanilla Ian Kendrich C
机构信息
Department of Health, Research Institute for Tropical Medicine, 9002 Research Drive, Filinvest Corporate City, Alabang, Muntinlupa City 1781, Philippines.
Institute of Biology, University of the Philippines Diliman, Quezon City 1101, Philippines.
出版信息
Trop Med Infect Dis. 2025 May 20;10(5):140. doi: 10.3390/tropicalmed10050140.
is the intermediate host of , the causative species of schistosomiasis in the Philippines. Conventionally, risk areas are identified by procedures requiring highly skilled personnel and constant surveillance efforts. Recent developments in disease diagnostics explore the utilization of environmental DNA as targets for polymerase chain reactions in disease surveillance. In this study, a low-cost, specific, and efficient SYBR Green-based real-time PCR assay to detect DNA from water samples was developed, optimized, and validated. Primers were designed based on the A18 microsatellite region of The assay exhibited a detection limit of one copy number per microliter at 99.4% efficiency and R = 0.999, which specifically amplified DNA only. Validation of this assay in environmental water samples demonstrated 100% PPV and NPV values, suggesting its potential as a tool for identifying risk areas, pathogen-directed surveillance, policy making, and disease control.
是菲律宾血吸虫病致病物种的中间宿主。传统上，风险区域是通过需要高技能人员和持续监测努力的程序来确定的。疾病诊断的最新进展探索了利用环境DNA作为疾病监测中聚合酶链反应的靶标。在本研究中，开发、优化并验证了一种低成本、特异性强且高效的基于SYBR Green的实时PCR检测方法，用于从水样中检测DNA。基于的A18微卫星区域设计了引物。该检测方法在效率为99.4%且R = 0.999时，检测限为每微升一个拷贝数，仅特异性扩增DNA。该检测方法在环境水样中的验证显示出100%的阳性预测值和阴性预测值，表明其作为识别风险区域、病原体定向监测、政策制定和疾病控制工具的潜力。
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