利用双抗夹心-酶联免疫吸附试验(DAS-ELISA)和反转录-聚合酶链式反应(RT- PCR)方法对来自韩国的番茄种子进行种传病毒检测.结果表明,在该批番茄种子中检测到番茄花叶病毒(ToMV)和烟草花叶病毒(TMV).PCR产物测序结果表明,ToMV特异引物(ToA/ToB)与TMV特异引物(TA/TB)扩增的片段均为ToMV的外壳蛋白(CP)基因及3′非编码区(3′-UTR),该片段与其他ToMV分离物的核苷酸序列相似性为98.2％～99.9％.本研究利用重新设计合成TMV和ToMV特异引物对该批种子进行RT-PCR检测,仅ToMV特异性引物(ToMfl/ToMr1)扩增到670 bp的预期片段,TMV特异引物(TMf1/TMr1)则未出现特异性扩增,表明重新设计的引物可准确区分ToMV和TMV.以上结果证实该批番茄种子仅携带ToMV.%The tomato Lycopersicon esculentum seeds imported from South Korea were detected with DAS-ELISA and RT-PCR methods. The results showed that both the Tobacco mosaic vinus(TMV)and the Tomato mosaic virus (ToMV) were found in the seed lots. Sequence determination proved that the fragment (703 bp) with ToMV-spe-cific primers (ToA/ToB) and the fragment (698 bp) with TMV-specific primers (TA/TB) were composed of coat protein gene and 3'-UTR of ToMV, sharing identities of 98.2%-99.9% with other ToMV isolates at nucleotide sequence level, which indicated that the previously published primer pair TA/TB was not suitable for diagnosis of TMV. In this study, the primers ToMf1/ToMrl and TMfl/TMrl were designed for discriminating between ToMV and TMV and validated on the ELISA positive samples. The expected RT-PCR products (670 bp) of ToMfl/ToMrl specific to ToMV were readily visible in the ELISA positive samples, while no target fragment (677 bp) of TMfl/TMrl specific to TMV was observed.
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