通过对β-微管蛋白（tub1）基因片段进行变性梯度凝胶电泳（DGGE）和扩增阻滞突变系统聚合酶链反应（ARMS-PCR）对长孢镰刀菌及相关物种进行特异性检测。
Specific detection of Fusarium langsethiae and related species by DGGE and ARMS-PCR of a beta-tubulin (tub1) gene fragment.
作者信息
Mach R L, Kullnig-Gradinger C M, Farnleitner A H, Reischer G, Adler A, Kubicek C P
机构信息
Institute for Chemical Engineering, Division of Applied Biochemistry and Gene Technology, l Vienna University of Technology, Getreidemarkt 9/1665, Vienna A-1060 Austria.
出版信息
Int J Food Microbiol. 2004 Sep 15;95(3):333-9. doi: 10.1016/j.ijfoodmicro.2003.12.011.
Fusarium langsethiae was recently described as a new toxigenic Fusarium species, which morphologically resembles Fusarium poae, but exhibits a mycotoxin pattern related to Fusarium sporotrichioides. To develop tools for early and specific detection of F. langsethiae and distinguishing it from related species of section Sporotrichiella and Discolor (F. poae, F. sporotrichioides, Fusarium kyushuense, Fusarium robustum, Fusarium sambucinum and Fusarium tumidum) sequence variations in their beta-tubulin-encoding (tub1) gene were employed to design two PCR-based methods, denaturing gradient gel electrophoresis (DGGE) and amplification refractory mutation system (ARMS)-PCR. DGGE reliably separated all these strains, even from mixtures and in the presence of DNA from their natural hosts Zea mais, Triticum aestivum and Avena sativa. In addition, a tetraprimer ARMS-PCR, which employs two primer pairs to amplify, respectively, two different fragments of tub1 in a single PCR reaction resulted in rapid differentiation between F. langsethiae, F. sporotrichioides and F. poae according to the number of amplicons (four, two and one, respectively). These two methods will thus be worthwhile tools in the specific detection of F. langsethiae in infected crops.
最近，朗氏镰刀菌被描述为一种新的产毒镰刀菌物种，其形态与禾谷镰刀菌相似，但表现出与拟分枝孢镰刀菌相关的霉菌毒素模式。为了开发早期特异性检测朗氏镰刀菌并将其与分枝孢镰刀菌组和变色镰刀菌组的相关物种（禾谷镰刀菌、拟分枝孢镰刀菌、九州镰刀菌、粗壮镰刀菌、接骨木镰刀菌和肿胀镰刀菌）区分开来的工具，利用它们β-微管蛋白编码（tub1）基因的序列变异设计了两种基于PCR的方法，变性梯度凝胶电泳（DGGE）和扩增阻滞突变系统（ARMS）-PCR。DGGE能够可靠地分离所有这些菌株，即使是从混合物中，以及在存在其天然宿主玉米、普通小麦和燕麦DNA的情况下。此外，一种四引物ARMS-PCR，在单个PCR反应中使用两对引物分别扩增tub1的两个不同片段，根据扩增子数量（分别为四个、两个和一个）实现了朗氏镰刀菌、拟分枝孢镰刀菌和禾谷镰刀菌之间的快速区分。因此，这两种方法将成为检测受感染作物中朗氏镰刀菌的有价值工具。
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