柑桔黄龙病环介导等温扩增检测方法建立及应用  

Rapid Detection of Citrus Huanglongbing Using a LAMP Method

作　　者：彭军[1] 廖孝文 杨海中 曾凡云[1] 龙海波[1] 裴月令[1] 郭建荣[1] 

机构地区：[1]中国热带农业科学院环境与植物保护研究所农业部热带作物有害生物综合治理重点实验室海南省热带农业有害生物监测与控制重点实验室,海南海口571101 [2]海南省琼中黎族苗族自治县农业技术推广中心,海南琼中572900

出　　处：《热带作物学报》2013年第10期1998-2003,共6页Chinese Journal of Tropical Crops

基　　金：中央级公益性科研院所基本科研业务费专项资金(中国热带农业科学院本级)项目(1630042012001&1630042013021);中国热带农业科学院环境与植物保护研究所引进人才科研启动基金项目(Hzs1201);海南省国际科技合作重点项目(2012-GH-007)

摘　　要：以柑桔黄龙病(Citrus Huanglongbing,HLB)病原菌16s rDNA保守区域为靶标设计LAMP引物,建立柑桔黄龙病的环介导等温核酸扩增(loop-mediated isothermal amplification assay,LAMP)检测技术。LAMP检测方法具有极高的检测特异性和灵敏性,其检测下限约为1 pg/μL质粒DNA,是PCR检测灵敏度的100倍,能快速、准确地对田间疑似样品进行检测。建立的柑桔黄龙病LAMP检测方法是对黄龙病检测方法的拓展和延伸,可以很好地应用于田间检测,可满足基层以及科研单位对该病害检测的需要。With the aim to develop a loop-mediated isothermal amplificatioan assay （LAMP） for rapid detection of Citrus Huanglongbing （HLB） caused by Candidatus liberibacter asisticus.The conserved 16S rDNA sequence published in GenBank （accession no.AY192576） was used to design a set of four special primers that recognized six distinct sequences of the conserved region of HLB genome.The detection limit of HLB-specific LAMP assay was about 1 pg/μL plasmid DNA,which was 100 times more sensitive than PCR method.The amplification products could be visual inspection by naked eyes with a closed-tube detection technique using SYBR Green Ⅰ staining.The positive products were changed to green,while the negative remain orange.The LAMP is a further extension of HLB detection method and it is a simple and rapid for detection of HLB in field samples,which would be used for field surveys and routine screening tissue culture materials for mass propagation.

关 键 词：柑桔黄龙病 环介导等温核酸扩增技术 检测 

分 类 号：S666.1[农业科学—果树学]