多重PCR技术检测4种大豆镰孢菌根腐病病原

镰孢菌是引起大豆根腐病的常见病原菌，传统检测病原镰孢菌的方法存在操作繁琐、时间长、成本高和 效率低等缺点，建立一种快速检测方法显得尤为重要。本研究基于翻译延伸因子序列（EF-1α）建立了检测镰孢菌 （Fusarium species）的多重PCR反应体系，检测了多重PCR体系灵敏度，模拟侵染样本验证多重PCR技术的可行性。 实验结果表明，该方法能够通过扩增片段的大小区分锐顶镰孢菌（Fusarium acuminatum）、尖孢镰刀菌（Fusarium oxysporum）、茄病镰孢菌（Fusarium solani）和禾谷镰孢菌（Fusarium graminearum）。灵敏度检测显示，最低检测基因 组DNA浓度达1×10-4 ng/μL；模拟侵染样本实验表明，使用镰孢菌和大豆基因组混合样本作为模板，在DNA浓度为 100 ng/μL时仍能准确检测出目的条带。因此。以翻译延伸因子序列为靶标建立的多重PCR技术，能够灵敏、特异 性地检测出该四种镰孢菌，可在复合侵染引起大豆镰孢菌根腐病的病原菌鉴定中准确检测。

Establishment of a multiplex PCR for detection of four Fusarium pathogens of soybean root rot disease

Fusariumspecies are the most common pathogens of soybean root rot disease. Traditional detection methods for Fusarium are complicated, time and money comsumming with low efficiency. It’s particularly important to establish a rapid detection method for detection of soybean Fusarium root rot disease. In our research, a multiplex PCR system was established based on the translation elongation factor gene(EF-1α). The sensitivity of the multiplex PCR reaction system and viability to detect simulating infection soybean tissues were carried out, the lowest concentration of genome DNA is 1×10^-4 ng/μL. We successfully detected Fusarium acuminutum, Fusarium oxysporum, Fusarium solani and Fusarium graminearum with the multiplex PCR detection system. The EF-1α bands were successfully amplified with genomes DNA of 100 ng/μL of the 4 Fusarium species or simulating infection samples as templates. In conclusion, a multiplex PCR detection method based on translation elongation factor gene was established which worked well to distinguish 4 different Fusarium species, the method could be used to diagnose Fusarium pathogens in multiple infection.