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根据GenBank登录的柑橘褪绿矮化相关病毒(Citrus chlorotic dwarf-associated virus,CCDaV)的移动蛋白(MP)基因序列设计了一组特异性引物,经体系优化,建立了CCDaV的环介导等温核酸扩增(LAMP)检测方法。结果显示该方法能特异扩增CCDaV,与其他5种柑橘病原物(柑橘衰退病毒、柑橘碎叶病毒、柑橘裂皮病类病毒、温州蜜柑萎缩病毒和柑橘黄龙病菌)不发生反应;灵敏度为PCR的100倍,与实时荧光PCR的灵敏度相同。用LAMP方法对50个田间样品进行检测,与PCR和实时荧光PCR的检测结果一致,证明该方法可用于田间样品的检测。该LAMP检测方法可特异、灵敏、快速地对CCDaV样品进行检测。
A set of specific primers was designed based on the sequence of the mobile protein (MP) gene of citrus chlorotic dwarf-associated virus (Citrus chlorotic dwarf-associated virus) registered in GenBank. After optimization of the system, a circular-mediated isothermal nucleic acid Amplification (LAMP) detection method. The results showed that this method could specifically amplify CCDaV and did not react with the other five citrus pathogens (Citrus recession virus, Citrus broken leaf virus, Citri distemper virus, Citrus wenzhou wilt virus and Citrus Huanglongbing); the sensitivity was PCR 100 times the same sensitivity as real-time fluorescence PCR. Fifty field samples were tested by LAMP method, which was consistent with PCR and real-time PCR. The results showed that the method was suitable for the field samples. The LAMP detection method can detect CCDaV samples specifically, sensitively and rapidly.