油菜茎基溃疡病菌LAMP-HNB检测方法的建立  

Establishment of LAMP-HNB method for detection of Leptosphaeria maculans in rape

作　　者：龙阳 马新华 袁俊杰 卢乃会 杨卓瑜 魏霜[2] 王卫芳[2] 

机构地区：[1]湛江出入境检验检疫局,广东湛江524042 [2]广东出入境检验检疫局,广东广州510623

出　　处：《广东农业科学》2017年第10期66-69,175,共5页Guangdong Agricultural Sciences

基　　金：广东出入境检验检疫局科技计划项目(2016GDK32)

摘　　要：油菜茎基溃疡病菌是油菜上最严重的真菌病害之一,是我国重要的检疫性有害生物。因此为防范其危害我国油菜产业生产安全,建立油菜茎基溃疡病菌的快速筛查方法具有重要意义。基于环介导等温扩增法(LAMP),以ITS(内转录间隔区)为靶序列并结合指示剂羟基萘酚蓝(HNB)建立了一种基于颜色判定的简单、快速和灵敏的油菜茎基溃疡病菌的快速检测方法,并对其特异性、灵敏性进行评价以及实际样品进行检测。结果显示,该方法在62℃等温条件下仅需80 min即可通过肉眼直接目测试验结果。LAMP-HNB法与标准PCR检测方法对比显示其具有良好的特异性以及更高的灵敏度,利用该方法对10批油菜籽样品进行检测,共检出5批阳性样品,与标准中PCR检测方法结果一致。该方法的建立为油菜茎基溃疡病菌的检疫鉴定提供了新途径。Leptosphaeria maculans is one of the most serious fungal diseases in rape. It is important to establish a rapid detection method to protect the safety of rapeseed industry in China. The loop-mediated isothermal amplification(LAMP) method was developed to detect L. maculans,and the hydroxy naphthol blue(HNB)was used as indicator. The specificity and sensitivity of the method were tested. The result can be obtained within 80 minutes at 62℃ and judged by the eye. The results showed that LAMP-HNB method had a higher sensitivity than conventional PCR. A total 5 positive samples for L. maculans were detected from 10 samples by the LAMP-HNB method,which was in accordance with the result by conventional PCR. LAMP-HNB is a new method to detect L. maculans.

关 键 词：油菜 茎基溃疡病菌 环介导等温扩增 羟基萘酚蓝 检疫 

分 类 号：S41-30[农业科学—植物保护]