3种葡萄座腔菌的检测方法比较
Comparison of Three Detection Methods for Botryosphaeria dothidea
王丽 1侯珲 2涂洪涛 1黄天祥 2柳友亮 2袁洪波1
作者信息
- 1. 中国农业科学院郑州果树研究所,郑州 450009;中国农业科学院中原研究中心,河南新乡 453003
- 2. 中国农业科学院郑州果树研究所,郑州 450009
- 折叠
摘要
为精准检测葡萄座腔菌(Botryosphaeria dothidea)引起的果树病害,基于B.dothidea的ITS序列设计常规PCR、巢式PCR和LAMP引物,对3种检测方法的灵敏度进行比较分析,依据研究结果选用巢式PCR和LAMP反应检测田间病样.结果表明,常规PCR、巢式PCR和LAMP检测方法均能特异性检测出B.dothidea.普通PCR检测到的DNA最低浓度为10-1 ng/μL,巢式PCR的检测限为104 ng/uL,LAMP方法的检测限为10-5ng/μL.灵敏度较高的巢式PCR和LAMP方法都可用于检测B.dothidea引起的田间病害样品.LAMP方法检测田间疑似病害样品的检出率为65.0％、常规PCR为58.8％、巢式PCR为62.5％,而常规分离鉴定方法的检出率仅为27.5％.
Abstract
Being able to rapidly and reliably detect Botryosphaeria dothidea will enable better disease monitoring and control efforts.In the present study,we developed conventional PCR,nested PCR,and LAMP assays in order to specifically differentiate between B.dothidea and other fungal species.Then,the sensitivity of the three detection methods was compared and analyzed.Finally,these assays were used to detect diseased samples collected from fields.The results showed that the conventional PCR,nested PCR and LAMP assays were able to detect and diagnose fruit diseases caused by B.dothidea.Sensitivity analyses revealed that the detection limit for conventional PCR assay was 10-1 ng/μL of B.dothidea DNA,whereas the nested PCR assay had a 10-4 ng/μL detection limit,and the LAMP assay was sensitive to as little as 10-5 ng/μL of fungal DNA.The nested PCR and LAMP assays can be reliably utilized to diagnose fruit diseased samples from field caused by B.dothidea.The positive rates of the LAMP detection,nested PCR,conventional PCR and traditional identification method were 65.0％,62.5％,58.8％and 27.5％respectively.
关键词
果树病害/葡萄座腔菌/聚合酶链式反应/环介导等温扩增技术/检测Key words
fruit diseases/Botryosphaeria dothidea/PCR/LAMP/detection引用本文复制引用
基金项目
河南省重点研发专项(221111111800)
中国农业科学院科技创新工程"猕猴桃资源与育种科研团队"(CAAS-ASTIP-2023-ZFRI)
长江猕猴桃产业技术研究中心(CJZX20210109)
出版年
2024
国家科技期刊平台
NSTL
万方数据