| 吕 燕1, 2, 郭立新1, 3, 段维军1, 3*.基于TaqMan MGB探针的可可花瘿病菌快速检测方法[J].植物保护,2022,48(5):220-226. |
| 基于TaqMan MGB探针的可可花瘿病菌快速检测方法 |
| Development of TaqMan MGB fluorescent real-time PCR assay for detection of Albonectria rigidiuscula |
| 投稿时间：2021-09-16 修订日期：2021-11-24 |
| DOI：10.16688/j.zwbh.2021509 |
| 中文关键词: 可可花瘿病菌 实时荧光PCR 检测 TaqMan MGB探针 |
| 英文关键词:Albonectria rigidiuscula real-time fluorescent PCR detection TaqMan MGB probe |
| 基金项目:国家重点研发计划(2022YFF0608804); 国家自然科学基金(NSFC31770009); 浙江省公益技术研究项目(LGF20C140001); 海关总署科研项目(2020HK162); 宁波市社会公益科研项目(2022S010) |
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| 中文摘要: |
| 可可花瘿病菌是一种我国进境植物检疫性真菌?本文根据可可花瘿病菌EF1α基因的保守序列, 设计并合成1对特异性的实时荧光PCR引物和1条TaqMan MGB探针, 建立了可可花瘿病菌的实时荧光PCR检测方法?特异性试验结果表明, 该检测方法能够特异性检出可可花瘿病菌; 实时荧光PCR优化反应条件为引物终浓度0.2 μmol/L, 探针终浓度0.6 μmol/L; 灵敏度试验结果表明, 20 μL反应体系中可可花瘿病菌DNA含量最低检测限为10 pg; 重复性试验结果表明, 该检测方法的重复性和稳定性良好; 接种试验样品检测结果表明, 该方法可用于疑似携带可可花瘿病菌样品的检测与初筛?本文建立的方法具有良好的灵敏性?特异性和应用性, 为可可花瘿病菌早期快速检测提供了一种有效手段? |
| 英文摘要: |
| Albonectria rigidiuscula is a quarantine fungus in the list of quarantine pests in China. A species-specific real-time polymerase chain reaction (qPCR) assay was developed for the detection of A. rigidiuscula. A pair of specific primers and a TaqMan MGB probe were designed and synthesized according to the conserved sequence of EF1α gene. The optimal primer concentration and probe concentration were 0.2 μmol/L and 0.6 μmol/L, respectively. The minimal detectable concentration of targeted DNA was 10 pg in 20 μL reaction mixture. The method had good repeatability and stability, and could be used for the detection and preliminary screening of the samples suspected of carrying A. rigidiuscula. The method was sensitive, specific and applicable, which provided a valuable tool for early rapid detection and identification of A. rigidiuscula. |
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