应用实时荧光定量RT-PCR高效检测葡萄病毒B
Effective detection of Grapevine virus B by real-time fluorescent quantitative RTPCR
查看参考文献16篇
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文摘
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本研究建立了葡萄病毒B的SYBR GreenⅠ实时荧光定量RT-PCR(RT-qPCR)检测技术。该技术标准曲线扩增效率102.4%,相关系数0.999,最低检测限达10-4倍稀释cDNA,灵敏度为常规RT-PCR的100倍。重复性试验组内和组间变异系数分别为0.00%~ 0.65%和0.02%~ 2.00%,表明检测稳定性好。该技术对田间葡萄样品检测适用范围广,对枝条和老叶柄检测效果最好,冬季枝条和春夏秋季所有老叶柄样品检出率均为100%,与常规RT-PCR检测结果一致。对于其他季节或部位样品,RT-qPCR检出率(43% ~ 74%)则普遍高于常规RT-PCR(5% ~ 71%),特别是春季样品和春夏秋季所有嫩叶样品,检出率比常规RT-PCR分别高31%和38%。对来自我国13个省21个品种的52份田间葡萄样品检测结果表明,RTqPCR共检测到6个样品为阳性,检出率11.5%,为常规RT-PCR(检出率5.8%)的2倍。 |
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其他语种文摘
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A SYBR GreenⅠRT-qPCR method for Grapevine virus B (GVB) was established.An excellent linear correlation(0.999) and amplification efficiency (102.4%) were obtained from standard curve.The detection limit of the method was 10-4dilution fold,which was 100 times higher than conventional RT-PCR.The coefficient of variation ranging from 0.00%-0.65% intra group and 0.02%-2.00% inter group indicated the excellent stability and reproducibility of the method.The RT-qPCR method could be used to detect a wide range of field sample types.The dormant branches and old petioles were the best materials for GVB detection,the detection rates of which were all 100% and were the same as conventional RT-PCR.However,for samples from other seasons and positions,the detection rates of RT-qPCR (43% to 74%) were generally higher than that of conventional RT-PCR (5% to 71%).Especially for the samples in spring and young leaves in each season,the detection rates of RT-qPCR were 31 and 38 percent higher than conventional RT-PCR,respectively.Six out of 52 samples (belonging to 21 cultivars) from 13 provinces in China were detected to be positive by RT-qPCR(11.5%),which was two times that of conventional RT-PCR (5.8%). |
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来源
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植物病理学报
,2019,49(4):569-576 【核心库】
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DOI
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10.13926/j.cnki.apps.000381
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关键词
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葡萄
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葡萄病毒B
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实时荧光定量RT-PCR
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常规RT-PCR
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地址
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中国农业科学院果树研究所,国家落叶果树脱毒中心, 兴城, 125100
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语种
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中文 |
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文献类型
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研究性论文 |
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ISSN
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0412-0914 |
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学科
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植物保护 |
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基金
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国家现代农业产业技术体系建设专项资金项目
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文献收藏号
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CSCD:6566495
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