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首页> 外文期刊>Plant Disease >Rapid and Sensitive Detection of Meloidogyne graminicola in Soil Using Conventional PCR, Loop-Mediated Isothermal Amplification, and Real-Time PCR Methods
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Rapid and Sensitive Detection of Meloidogyne graminicola in Soil Using Conventional PCR, Loop-Mediated Isothermal Amplification, and Real-Time PCR Methods

机译：使用常规PCR，环介导的等温扩增和实时PCR方法快速和灵敏地检测土壤中的Meloidogyne Graminicola

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摘要

Meloidogyne graminicola is one of the major plant-parasitic nematodes (PPNs) that affect rice agriculture. Rapid identification and quantification of M. graminicola in soil is crucial for early diagnosis so that measures can be taken to reduce the impact of PPN diseases and ensure food security. In this study, M. graminicola species-specific primers for conventional PCR, loop-mediated isothermal amplification (LAMP), and real-time PCR were designed based on the sequence-characterized amplified region. The primers were highly specific and sensitive, and only samples containing M. graminicola DNA showed positive results. The sensitivity of LAMP and real-time PCR (two second-stage juvenile [J2] M. graminicola in 100 g of soil) was higher than that of conventional PCR (200 J2s in 100 g of soil). A standard curve (correlation coefficient R-2 = 0.970, P < 0.001) was generated by amplifying DNA extracted from 0.5 g of soil, and a significant correlation was observed between the number of M. graminicola determined by microscopic examination and that predicted from the standard curve (R-2 = 0.477, P = 0.0160). In quantification analyses of M. graminicola isolated from 31 naturally infested soils, the sensitivity of LAMP and real-time PCR (22 M. graminicola in 100 g of soil) was higher than that of conventional PCR (211 M. graminicola in 100 g of soil). The conventional PCR, LAMP, and real-time PCR methods have the potential to provide a useful platform for rapid species identification according to the experimental conditions. The real-time PCR assay and standard curve can be used for quantification of M. graminicola. These newly developed assays will help to facilitate the control of these economically important PPNs.
机译：禾谷根结线虫是影响水稻农业生产的主要植物寄生线虫之一。快速鉴定和量化土壤中的格拉米菌对早期诊断至关重要，以便采取措施减少PPN疾病的影响，确保粮食安全。在本研究中，基于扩增区的序列特征，设计了用于常规PCR、环介导等温扩增（LAMP）和实时PCR的格拉霉种特异性引物。该引物具有高度特异性和敏感性，只有含有格拉米菌DNA的样本显示阳性结果。LAMP和实时PCR（100g土壤中的两个二级幼虫[J2]禾谷霉）的灵敏度高于常规PCR（100g土壤中的200J2s）。通过放大从0.5 g土壤中提取的DNA，生成标准曲线（相关系数R-2=0.970，P<0.001），并观察到显微镜检查确定的格拉米菌数量与标准曲线预测的数量之间存在显著相关性（R-2=0.477，P=0.0160）。在对从31个自然感染土壤中分离出的禾本科菌进行定量分析时，LAMP和实时PCR（100 g土壤中22株禾本科菌）的灵敏度高于常规PCR（100 g土壤中211株禾本科菌）。传统的PCR、LAMP和实时PCR方法有可能根据实验条件为快速物种鉴定提供有用的平台。实时PCR检测和标准曲线可用于格拉米菌的定量。这些新开发的分析方法将有助于促进这些经济上重要的PPN的控制。

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  • 来源
    《Plant Disease》 |2021年第2期|共8页
  • 作者

    He Qingcong; Wang Dongwei; Tang Bei; Wang Jian; Zhang Deyong; Liu Yong; Cheng Feixue;

  • 作者单位

    Hunan Acad Agr Sci Inst Plant Protect Key Lab Integrated Management Pests &

    Dis Hort Cr Changsha 410125 Peoples R China;

    Hunan Acad Agr Sci Inst Plant Protect Key Lab Integrated Management Pests &

    Dis Hort Cr Changsha 410125 Peoples R China;

    Hunan Acad Agr Sci Inst Plant Protect Key Lab Integrated Management Pests &

    Dis Hort Cr Changsha 410125 Peoples R China;

    Hunan Acad Agr Sci Agr Econ &

    Reg Planning Res Inst Changsha 410125 Peoples R China;

    Hunan Acad Agr Sci Inst Plant Protect Key Lab Integrated Management Pests &

    Dis Hort Cr Changsha 410125 Peoples R China;

    Hunan Acad Agr Sci Inst Plant Protect Key Lab Integrated Management Pests &

    Dis Hort Cr Changsha 410125 Peoples R China;

    Hunan Acad Agr Sci Inst Plant Protect Key Lab Integrated Management Pests &

    Dis Hort Cr Changsha 410125 Peoples R China;

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  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 植物保护;
  • 关键词

    conventional PCR; loop-mediated isothermal amplification (LAMP); Meloidogyne graminicola; real-time PCR; sensitivity; specificity;

    机译：常规PCR;环介导的等温扩增（灯）;Meloidogyne Graminicola;实时PCR;灵敏度;特异性;

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