, Clinglobal, B03/04, The Tamarin Commercial Hub, Jacaranda Avenue, Tamarin, 90903, Mauritius.
, Clinomics, Uitzich Road, Bainsvlei, Bloemfontein, 9338, South Africa.
Sci Rep. 2024 Jun 12;14(1):13584. doi: 10.1038/s41598-024-64401-0.
A novel, turnkey, field-based workflow was developed and validated using Rhipicephalus microplus DNA as a template to detect the presence of the voltage-gated sodium channel kdr mutation. The field-based compatible workflow comprises manual sample homogenization for DNA extraction, PCR amplification of the targets in a closed tube, and end-point detection of the PCR products. An R. microplus species-specific assay was also included to confirm species identity and ensure the validity of the kdr mutation assay. The assays were sensitive and specific to the targets, and the workflow resulted in a turnaround time of approximately 1 h at a low cost. The novel combination of PCR with closed-tube and end-point fluorescent detection allows for easy conversion of existing conventional lab-based PCR assays into field-based detection assays. The incorporation of custom-designed 3D-printed components in the workflow provides easy adaptability and modification of the components for diverse nucleic acid detection workflows.
开发并验证了一种新颖的、交钥匙型、基于现场的工作流程，该流程使用 Rhipicephalus microplus DNA 作为模板来检测电压门控钠离子通道 kdr 突变的存在。基于现场的兼容工作流程包括手动样品均质化以提取 DNA、在封闭管中对靶标进行 PCR 扩增，以及对 PCR 产物进行终点检测。还包括了一种 R. microplus 种特异性检测，以确认物种身份并确保 kdr 突变检测的有效性。这些检测对靶标具有敏感性和特异性，并且该工作流程的周转时间约为 1 小时，成本低。PCR 与封闭管和终点荧光检测的新颖组合允许将现有的常规实验室基于 PCR 检测转化为基于现场的检测。工作流程中包含定制设计的 3D 打印组件，可轻松适应和修改组件，以适应不同的核酸检测工作流程。
