向日葵茎溃疡病菌数字PCR检测方法的建立
Development of digital PCR detection method for Diaporthe helianthi on sunflower
辛淑香 1蒋刚强 2龙志新 2张小菊 2田沁怡 2罗明 3王翀 2虎子辉 2史博 2翟俊丽4
作者信息
- 1. 新疆农业大学农学院 新疆乌鲁木齐 830052;乌鲁木齐海关
- 2. 乌鲁木齐海关
- 3. 新疆农业大学农学院 新疆乌鲁木齐 830052
- 4. 卡拉苏海关
- 折叠
摘要
本研究采用国家标准GB/T 31792-2015《向日葵茎溃疡病菌检疫鉴定方法》的引物探针,通过构建阳性质粒、测试其特异性、灵敏度和重复性,建立了向日葵茎溃疡病菌的数字PCR检测方法.测试结果表明:引物和探针对供试的4株向日葵茎溃疡病菌有特异性扩增,其他14株病菌无扩增反应;本试验方法最低检出限为0.008 pg/pL;重复性试验RSD值＜25％,说明具有良好的重复性.通过对收集的14份样品进行检测,未检出向日葵茎溃疡病菌.因此,该方法具有特异性强、灵敏度高和稳定的重复性等特点,为向日葵茎溃疡病菌早期监测和预警提供了可靠有效的技术支撑.
Abstract
In this study,the primer-probe of national standard GB/T 31792-2015"Quarantine and Identifi-cation Method of Sunflower Stem Ulcer Pathogen"was used,and the digital PCR detection technology of Diaporthe helianthi was established by constructing positive plasmid and testing its specificity,sensitivity,and repeatability.The test results showed that the primers and probes had specific amplification for 4 strains of Diaporthe helianthi,while the other 14 strains had no amplification reaction.The minimum de-tection limit of this test method is 0.008 pg/µL;The RSD of the repeatability test was less than 25％,which shows that it has good repeatability.By detecting 14 samples collected,no Diaporthe helianthi was detected.Therefore,this method has strong specificity,high sensitivity,and stable repeatability,providing re-liable and effective technical support for early monitoring and early warning of Diaporthe helianthi.
关键词
向日葵茎溃疡病菌/数字PCR/绝对定量/检测Key words
Diaporthe helianthi/digital PCR/absolute quantification/detection引用本文复制引用
基金项目
新疆维吾尔自治区自然科学基金资助项目(2022D01A25)
海关总署科研计划项目(2023HK017)
出版年
2024
NETL
NSTL
万方数据