Rapid detection of Macrophomina phaseolina in common bean seeds using a visual loop-mediated isothermal amplification assay
Rapid detection of Macrophomina phaseolina in common bean seeds using a visual loop-mediated isothermal amplification assay
复制标题
使用视觉环介导等温扩增测定法快速检测普通菜豆种子中的菜豆大孔菌
DOI：
10.1007/s13313-017-0477-0
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发表时间：
2017
影响因子：
1.4
通讯作者：
S. Petrofeza
中科院分区：
文献类型：
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作者：
D. C. Rocha;M. B. Oliveira;Marcos Augusto de Freitas;S. Petrofeza
菜豆茎点霉Macrophomina phaseolina（Tassi）Goid是一种危害菜豆的病原菌，严重危害菜豆的生产，造成严重的经济损失。（common bean）.在这里，我们开发了一种快速，直观，灵敏的环介导等温扩增（LAMP）检测M。基于序列特征的扩增区域的菜豆种子中菜豆蛋白检测（GU018142.1）。在65 °C下45 min内获得了理想的检测条件。目视检查LAMP测定中羟基萘酚蓝染料的添加。在环境光下，阳性反应变成天蓝色，而阴性对照保持紫色。检测限为1皮克M。与其他5个种传病原体分离的DNA无交叉反应。LAMP检测结果表明，两种菜豆种子均被M.菜豆和人工感染病原体的种子。检测限为每批400粒种子中一粒受感染的种子。这种检测方法是可能的，没有事先提取真菌DNA，浸泡种子2-6小时。这些结果表明，LAMP检测提供了一个快速的定性诊断工具，检测M。菜豆种子
Macrophomina phaseolina(Tassi) Goid is a pathogen that causes significant economic losses by infestingPhaseolus vulgarisL. (common bean). Here, we developed a rapid, visual, and sensitive loop-mediated isothermal amplification (LAMP) assay forM. phaseolinadetection in common bean seeds based on sequence-characterized amplified regions (GU018142.1). The ideal conditions for detection were obtained in 45 min at 65 °C. The LAMP assay was visually inspected for the addition of hydroxynaphthol blue dye. Positive reactions turned sky blue while the negative controls remained violet under ambient light. The detection limit was 1 pg ofM. phaseolinagenomic DNA per reaction, and no cross reactions were observed with DNA isolated from five other seedborne pathogens. The LAMP assay detected both common bean seeds naturally infected byM. phaseolina, and seeds artificially contaminated with the pathogen. The limit of detection was one infected seed per lot of 400 seeds. This detection method was possible without prior fungal DNA extraction, by soaking seeds 2–6 h. These results suggest that the LAMP assay provides a rapid qualitative diagnostic tool for detectingM. phaseolinain bean seeds.
影响因子：
--
作者：
Kaneko, Hisatoshi;Kawana, Takashi;Suzutani, Tatsuo
通讯作者：
Suzutani, Tatsuo
影响因子：
2.7
作者：
Goto, Motoki;Honda, Eiichi;Hanaki, Ken-Ichi
通讯作者：
Hanaki, Ken-Ichi