Wetzel T, Candresse T, Ravelonandro M, Dunez J
Station de Pathologie Végétale, INRA, Villenave d'Ornon, France.
J Virol Methods. 1991 Aug;33(3):355-65. doi: 10.1016/0166-0934(91)90035-x.
A sensitive, polyvalent assay based on the polymerase chain reaction (PCR) was developed for plum pox potyvirus (PPV) detection. This technique was adapted for a single tube, the chemical denaturation and reverse transcription of the viral RNA followed by the PCR reaction yielding a 243-base-pair product. As few as 10 fg of purified viral RNA, corresponding to approximately 2000 viral particles, were detected in plant extracts. All PPV isolates tested were amplified, and the amplified fragments were analysed by restriction endonuclease digestion. An RsaI restriction site polymorphism in the amplified fragments allowed the discrimination of two groups of isolates. In a field indexing trial, the PCR assay proved to be more sensitive than molecular hybridization using 32P-labelled RNA probes for PPV detection.
开发了一种基于聚合酶链反应（PCR）的灵敏多价检测方法用于李痘病毒（PPV）的检测。该技术适用于单管操作，病毒RNA经化学变性和逆转录后进行PCR反应，产生一个243碱基对的产物。在植物提取物中可检测到低至10 fg的纯化病毒RNA，这大约相当于2000个病毒粒子。所有测试的PPV分离株均能被扩增，扩增片段通过限制性内切酶消化进行分析。扩增片段中的RsaI限制性酶切位点多态性可区分两组分离株。在田间检测试验中，PCR检测方法被证明比使用32P标记的RNA探针进行分子杂交检测PPV更为灵敏。
