兰花褐斑病菌可视化基因芯片检测技术
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摘要
兰花褐斑病菌(Acidovorax cattleyae,Ac)是一种严重为害兰花的检疫性有害生物。根据Lep A基因序列设计特异性引物和探针,利用可视化基因芯片技术,建立了Ac的基因芯片检测方法。测试结果显示,供试的3株兰花褐斑病菌菌株均为阳性,其他19株对照菌株为阴性。菌体DNA的检测灵敏度可达到450 fg,是常规PCR的100倍;人工模拟感染兰花样品的检测灵敏度为1.6×10~3 CFU。可视化基因芯片检测技术的强特异性,高灵敏度,易操作性和低成本特点适合口岸一线和基层实验室开展兰花褐斑病菌的快速检测。
Acidovorax cattleyae(Ac) is a quarantine pest of China which is seriously harmful to orchid.This paper reports the method of detecting Ac using visual gene chip technology. According to Lep A gene sequence we design specific primers and probes,the results of the detection of the three collected strains were positive,and the results of 19 other control strains were negative. The sensitivity of the detection bacteria DNA can reach 450 fg,which is 100 times that of conventional PCR,and the maximum sensitivity of the simulatively infected orchid leaf is 1.6 ×10~3 CFU. The visualization gene chip has the advantages of specificity,high sensitivity,easy operation,low requirement of equipment. Therefore,the Ac visualization gene chip technology can be effectively used for the detection of orchidaria.
Acidovorax cattleyae(Ac) is a quarantine pest of China which is seriously harmful to orchid.This paper reports the method of detecting Ac using visual gene chip technology. According to Lep A gene sequence we design specific primers and probes,the results of the detection of the three collected strains were positive,and the results of 19 other control strains were negative. The sensitivity of the detection bacteria DNA can reach 450 fg,which is 100 times that of conventional PCR,and the maximum sensitivity of the simulatively infected orchid leaf is 1.6 ×10~3 CFU. The visualization gene chip has the advantages of specificity,high sensitivity,easy operation,low requirement of equipment. Therefore,the Ac visualization gene chip technology can be effectively used for the detection of orchidaria.
引文
[1]冯洁.植物病原细菌分类最新进展.中国农业科学,2017,50(12):2305-2314.
[2]王春林等.潜在的植物检疫性有害生物图鉴.北京:中国农业出版社,2005:264-266.
[3]寸东义,赵文军,杜宇,等.进境兰花褐斑病菌的分离及PCR鉴定.植物检疫,2007,21(4):210-212.
[4]丁翠珍,赵文军,朱水芳,等.兰花褐斑病菌实时荧光PCR检测.植物病理学报,2010,40(3):235-241.
[5]谭建锡,周慧平,彭梓,等.兰花5种病毒可视化基因芯片检测方法建立.植物病理学报,2016(5):1-12.
[6]张靓,田茜,刘凤权,等.六种重要瓜类种传细菌可视基因芯片筛查方法研究.农业生物技术学报,2013(1):120-123.
[7]徐文华,范燕萍.蝴蝶兰细菌性褐斑病病原菌初步鉴定.植物保护,2006,32(5):87-89.
[8]丁爱东,刘朝祯,戚佩坤.蝴蝶兰细菌性褐斑病鉴定.华南农业大学学报,1993,14(4):124-126.
[9]蔡孟庭.Pectobacterium chrysanthem病原性相关基因之分析[D].台湾:中华医事科技大学,2013.
[10]廖太林,吴新华,李百胜,等.主要贸易国家对兰花的检疫要求.植物检疫,2007,21(5):325-326.
[2]王春林等.潜在的植物检疫性有害生物图鉴.北京:中国农业出版社,2005:264-266.
[3]寸东义,赵文军,杜宇,等.进境兰花褐斑病菌的分离及PCR鉴定.植物检疫,2007,21(4):210-212.
[4]丁翠珍,赵文军,朱水芳,等.兰花褐斑病菌实时荧光PCR检测.植物病理学报,2010,40(3):235-241.
[5]谭建锡,周慧平,彭梓,等.兰花5种病毒可视化基因芯片检测方法建立.植物病理学报,2016(5):1-12.
[6]张靓,田茜,刘凤权,等.六种重要瓜类种传细菌可视基因芯片筛查方法研究.农业生物技术学报,2013(1):120-123.
[7]徐文华,范燕萍.蝴蝶兰细菌性褐斑病病原菌初步鉴定.植物保护,2006,32(5):87-89.
[8]丁爱东,刘朝祯,戚佩坤.蝴蝶兰细菌性褐斑病鉴定.华南农业大学学报,1993,14(4):124-126.
[9]蔡孟庭.Pectobacterium chrysanthem病原性相关基因之分析[D].台湾:中华医事科技大学,2013.
[10]廖太林,吴新华,李百胜,等.主要贸易国家对兰花的检疫要求.植物检疫,2007,21(5):325-326.