| 单长林，周 圆，李孝军，杨赛军，邵炜冬.RT-LAMP 及RT-PCR 方法快速检测玉米褪绿斑驳病毒的比较与应用[J].广东农业科学,2017,44(7):83-90 |
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RT-LAMP 及RT-PCR 方法快速检测玉米褪绿斑驳病毒的比较与应用 |
| Comparison and application of RT-LAMP and RT-PCR methods for MCMV detection |
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| DOI：10.16768/j.issn.1004-874X.2017.07.014 |
| 中文关键词: 玉米褪绿斑驳病毒 反转录环介导等温扩增方法 RT-PCR 检测 |
| 英文关键词: MCMV RT-LAMP RT-PCR detection |
| 基金项目:舟山市科技计划项目（2015C31051） |
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| 中文摘要: |
| 利用玉米褪绿斑驳病毒外壳蛋白基因为靶标序列设计引物，通过优化反应温度、引物浓度、反
应时间以及评价环引物效果，建立了检测玉米褪绿斑驳病毒反转录环介导等温扩增（ RT-LAMP） 方法，并
与RT-PCR 方法比较。结果表明，RT-LAMP 在64℃等温条件下反应45 min，最低可检测到280 fg 的目的片
段，灵敏度是RT-PCR 的100 倍。以甘蔗花叶病毒、玉米矮花叶病毒、小麦线条花叶病毒、玉米白线花叶病
毒及玉米褪绿斑驳病毒等5 种病毒为检测对象，证明该方法针对玉米褪绿斑驳病毒具有很强的特异性。利
用RT-LAMP 方法对100 份模拟病毒感染的玉米种子进行检测，病毒检出率为100%。 |
| 英文摘要: |
| For the detection of Maize chlorotic mottle virus（MCMV），a reverse-transcription，loop mediated
isothermal amplification（ RT-LAMP） assay with high sensitivity and specificity was established. A set of five primers
were designed，based on the coat-protein sequence of MCMV. By optimizing the concentration of primers，the best
temperature，the suitable reaction time，and loop primer，the RT-LAMP was built by incubation at 64℃ for only 45
min with loop primer. The detection limit of RT-LAMP assay was 280 fg of MCMV RNA，which was 100 times more
sensitive than that of RT- PCR assay. High species-specificity of RT- LAMP method was confirmed by the assay of
5 pathogens such as MCMV，SCMV，MDMV， WSMV and MWLMV. In addition，all the simulated samples were
detected by LAMP assay，detection rate was 100%. |
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