| 毛宇宁, 梁鹏, 刘文波, 林春花, 缪卫国*, 郑服丛*.橡胶树白粉病菌分子检测技术的建立[J].植物保护,2016,42(4):119-123. |
| 橡胶树白粉病菌分子检测技术的建立 |
| PCR and nested-PCR for the molecular detection of Oidium heveae Steinm. |
| 投稿时间：2015-10-30 修订日期：2016-03-03 |
| DOI： |
| 中文关键词: 橡胶树白粉病菌 潜育期 分子检测 |
| 英文关键词:Oidium heveae incubation period molecular detection |
| 基金项目:国家“973”计划项目(2011CB111612); 国家现代农业产业技术体系(CARS-34-GW8); 国家自然科学基金(31160359); 教育部博士点基金(20104601110004); 海南大学科研启动资金(kyqd1006) |
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| 中文摘要: |
| 根据橡胶树白粉菌(Oidium heveae Steinm.)基因组中的特有保守序列OHS, 设计两对特异性引物OHF1/OHR1和OHF2/OHR2。以不同地区收集的6份O. heveae（OH1~OH6）和橡胶树胶孢炭疽病菌(Colletotrichum gloeosporioides Penz.)等5种非靶标病原菌及健康橡胶树叶片基因组DNA为模板, 建立了橡胶树白粉菌PCR及nested-PCR分子检测技术, 并验证了检测体系的特异性和灵敏度。结果表明, 引物OHF1/OHR1和OHF2/OHR2对橡胶树白粉菌均具有较高的特异性和灵敏度。其中引物OHF2/OHR2能检测到10 pg/μL的橡胶树白粉菌DNA, 而以OHF1/OHR1和OHF2/OHR2组合进行nested-PCR, 其最低DNA检测浓度达到0.01 fg/μL。人工接种试验中, 当孢子接种量为2×103个/叶时, PCR和nested-PCR检测体系可分别在接种4 d和24 h后检测到目的条带。表明nested-PCR对在叶片组织中处于潜育期的橡胶树白粉菌的检测更有效, 可为橡胶树白粉菌的检测提供一种简便而准确的检测方法。 |
| 英文摘要: |
| To establish methods for detection of powdery mildew of rubber tree by using conventional PCR and nested-PCR, two sets of specific primers OHF1/OHR1 and OHF2/OHR2 were designed respectively based on unique conserved sequence OHS of Oidium heveae. Six O. heveae samples (OH1-OH6) from different region, five strains of non-target pathogenic fungi and healthy rubber tree leaves were used to evaluate the specificity and sensitivity of conventional PCR and nested-PCR detection system. The results revealed that both OHF1/OHR1 and OHF2/OHR2 have high specificity and sensitivity. The detection limit is 10 pg/μL and 0.01 fg/μL DNA, respectively for conventional PCR (using primer OHF2/OHR2) and nested-PCR (using OHF1/OHR1 and OHF2/OHR2). Artificial inoculation test of O. heveae on rubber tree indicated that target band can be detected by PCR and nested-PCR at 4th day and 24 h after inoculation, respectively. These results suggested that the nested-PCR is more efficient than the conventional PCR for O. heveae in incubation period and provide an easy and accurate way for O. heveae detection. |
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