Division of Microbiology, National Institute of Health Sciences, Kamiyoga 1-18-1, Setagaya-ku, Tokyo 158-8501, Japan.
J Food Prot. 2010 Jun;73(6):1077-84. doi: 10.4315/0362-028x-73.6.1077.
To identify a rapid method for extracting a large amount of DNA from fungi associated with food hygiene, extraction methods were compared using fungal pellets formed rapidly in liquid media. Combinations of physical and chemical methods or commercial kits were evaluated with 3 species of yeast, 10 species of ascomycetous molds, and 4 species of zygomycetous molds. Bead grinding was the physical method, followed by chemical methods involving sodium dodecyl sulfate (SDS), cetyl trimethyl ammonium bromide (CTAB), and benzyl chloride and two commercial kits. Quantity was calculated by UV absorbance at 260 nm, quality was determined by the ratio of UV absorbance at 260 and 280 nm, and gene amplifications and electrophoresis profiles of whole genomes were analyzed. Bead grinding with the SDS method was the most effective for DNA extraction for yeasts and ascomycetous molds, and bead grinding with the CTAB method was most effective with zygomycetous molds. For both groups of molds, bead grinding with the CTAB method was the best approach for DNA extraction. Because this combination also is relatively effective for yeasts, it can be used to extract a large amount of DNA from a wide range of fungi. The DNA extraction methods are useful for developing gene indexes to identify fungi with molecular techniques, such as DNA fingerprinting.
为了鉴定一种从与食品卫生相关的真菌中快速提取大量 DNA 的方法，我们比较了利用液体培养基中快速形成的真菌菌球的提取方法。使用 3 种酵母、10 种子囊菌和 4 种接合菌评估了物理和化学方法的组合或商业试剂盒。珠磨是物理方法，其次是涉及十二烷基硫酸钠（SDS）、十六烷基三甲基溴化铵（CTAB）和氯化苄的化学方法，以及两种商业试剂盒。通过 260nm 的紫外吸光度计算数量，通过 260nm 和 280nm 的紫外吸光度比值确定质量，并分析全基因组的基因扩增和电泳图谱。对于酵母和子囊菌，SDS 法的珠磨是提取 DNA 最有效的方法，而 CTAB 法的珠磨对于接合菌最有效。对于这两组真菌，CTAB 法的珠磨是提取 DNA 的最佳方法。由于这种组合对于酵母也相对有效，因此可以用于从广泛的真菌中提取大量 DNA。这些 DNA 提取方法可用于开发基于分子技术（如 DNA 指纹图谱）鉴定真菌的基因指标。
