Potato spindle tuber viroid (PSTVd) is an important pathogen of potato infecting leaves, tubers as well as true potato seeds. We developed and validated a reverse transcription-loop mediated isothermal amplification (RT-LAMP) platform for visual, rapid, specific, and sensitive detection of PSTVd. The optimization of RT-LAMP reaction conditions and reagents concentrations were carried out with time, temperature, MgSO₄, dNTPs, and WarmStart Bst 2.0 polymerase. The assay is very simple and rapid which provide detectable results in less than 50 min by incubating all the reagents at 65 °C. The detection of RT-LAMP results could be accomplished as ladder-like bands in an agarose gel or visualized by naked eyes with inclusion of a SYBR Green I dye. The target specificity of RT-LAMP primers was assessed with an introduction of a simple restriction digestion step after the RT-LAMP assay. The RT-LAMP assay demonstrated absence of any cross-reaction with other, potato viruses. The analytical sensitivity of the assay was greater than RT-PCR either the test template was plasmid or total RNA. The detection of RT-LAMP was validated in potato samples and compared with real time PCR and RT-PCR. The RT-LAMP assay developed in this study has the potential as a beneficial tool in detection of PSTVd in a low-cost laboratory due to its simplicity, rapidness, and application in large samples.

马铃薯纺锤块茎类病毒（PSTVd）是马铃薯感染叶片，块茎以及真正的马铃薯种子的重要病原体。我们开发并验证了逆转录环介导的等温扩增（RT-LAMP）平台，用于PSTVd的视觉，快速，特异性和灵敏检测。RT-LAMP反应条件和试剂浓度的优化是随时间，温度，MgSO ₄，dNTPs和WarmStart Bst进行的2.0聚合酶。该测定非常简单且快速，通过在65°C下孵育所有试剂，可在不到50分钟的时间内提供可检测的结果。RT-LAMP结果的检测可以在琼脂糖凝胶中以梯状条带的形式完成，也可以通过肉眼观察并结合SYBR Green I染料来实现。RT-LAMP引物的靶标特异性通过在RT-LAMP测定后引入简单的限制性酶切消化步骤进行评估。RT-LAMP分析表明与其他马铃薯病毒没有任何交叉反应。无论测试模板是质粒还是总RNA，该测定的分析灵敏度均高于RT-PCR。RT-LAMP的检测已在马铃薯样品中得到验证，并与实时PCR和RT-PCR进行了比较。