Division of Plant Protection, ICAR-Central Potato Research Institute, Shimla, Himachal Pradesh, 171001, India; Center on Rabi Sorghum, ICAR-Indian Institute of Millets Research, Regional Station, Solapur, Maharashtra, 413006, India.
Division of Plant Protection, ICAR-Central Potato Research Institute, Shimla, Himachal Pradesh, 171001, India.
Virology. 2023 Oct;587:109872. doi: 10.1016/j.virol.2023.109872. Epub 2023 Aug 26.
Reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for detection of groundnut bud necrosis virus (GBNV) causing potato stem necrosis disease. The isothermal temperatures, reaction periods and concentrations of reaction mixture were optimized where, the assay worked well at 65 °C for 50 min, 6 U of WarmStart Bst 2.0 DNA polymerase, 1.4 mM dNTPs and 2.0 mM MgSO The optimized assay proved to be specific to GBNV with no cross reactivity to other viruses infecting potato in India. The specificity of RT-LAMP assay was found to be 100 fold more sensitive than that of RT-PCR. The developed assay was applied for the detection of GBNV from 80 potato leaf samples where 24 samples were found infected which was confirmed by RT-PCR. It was concluded that the RT-LAMP assay developed for detection of GBNV was specific, sensitive and suitable for its use in virus indexing under potato seed production programme.
逆转录环介导等温扩增（RT-LAMP）检测方法被开发出来，用于检测导致马铃薯茎坏死病的花生芽坏死病毒（GBNV）。优化了等温温度、反应时间和反应混合物浓度，在 65°C 下反应 50 分钟、6 U WarmStart Bst 2.0 DNA 聚合酶、1.4mM dNTPs 和 2.0mM MgSO4 的条件下，该检测方法效果良好。优化后的检测方法特异性针对 GBNV，与印度感染马铃薯的其他病毒无交叉反应。与 RT-PCR 相比，RT-LAMP 检测方法的特异性高 100 倍。该方法应用于检测 80 个马铃薯叶片样本中的 GBNV，其中 24 个样本被 RT-PCR 检测为阳性。研究结果表明，该方法特异性强、灵敏度高，适用于马铃薯种子生产计划中的病毒索引。
