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从一批进境的美国苜蓿草样品中分离得到了一株与苜蓿黄萎病菌(Verticillium albo-atrum)相似的分离物M1。该分离物原始菌落为白色,边缘规则呈圆形,菌丝较致密,气生菌丝较少,苜蓿组织块不被菌丝覆盖,菌落生长速度为小于2.5 mm/d;M1在PDA上进行纯培养,前6d内,菌落为白色圆形,容易产生分子孢子轮枝状分生孢子梗,7d后菌落中央表面因产生休眠菌丝开始变成黑褐色至黑色,20d后菌落的表面和背面大部分均变黑色,仍不产生微菌核和厚垣孢子。M1的DNA用V.albo-atrum特异引物Vaa1/Vaa2进行检测,PCR扩增后得到预期330 bp的产物片段,产物序列与V.albo-atrum相应序列的相似性为100%。该分离物接种苜蓿草根部,15d后引起苜蓿黄萎病的典型症状。根据分离物的形态特征、PCR检测结果、PCR产物序列分析,以及致病性测定结果,将进境美国苜蓿草样品中的分离物M1鉴定为苜蓿黄萎病菌。
A isolate M1 similar to Verticillium albo-atrum was isolated from a group of imported alfalfa samples. The original colonies of the isolates were white, the edges were round, the mycelium was dense, the number of aerial mycelia was less, the alfalfa tissue was not covered by mycelium, the colony growth rate was less than 2.5 mm / d; M1 was carried on PDA Pure culture, the first 6d, the colony is white round, easy to produce round spores of conidiospores stem spores, 7d after the central surface of the colony began to become dormant mycelium dark brown to black, 20d after the colony surface and back Most are black, still does not produce micro-bacteria and chlamydospores. The DNA of M1 was detected by Vaal / Vaa2 specific primers Val1 / Vaa2. After PCR amplification, the expected fragment of 330 bp was obtained. The similarity between the product sequence and V.albo-atrum was 100%. The isolate was inoculated with alfalfa roots and caused typical symptoms of alfalfa wilt after 15 days. According to the morphological characteristics of the isolates, PCR test results, PCR product sequence analysis and pathogenicity test results, M1, which entered the American alfalfa samples, was identified as Verticillium dahliae.