Rodrigues P, Venâncio A, Lima N
Centro de Investigação de Montanha (CIMO), ESA, Instituto Politécnico de Bragança, Campus de Santa Apolónia, Bragança, Portugal.
Centre for Biological Engineering, Universidade do Minho, Braga, Portugal.
Lett Appl Microbiol. 2018 Jan;66(1):32-37. doi: 10.1111/lam.12822. Epub 2017 Dec 11.
The aim of this work was to evaluate a fungal DNA extraction procedure with the lowest inputs in terms of time as well as of expensive and toxic chemicals, but able to consistently produce genomic DNA of good quality for PCR purposes. Two types of fungal biological material were tested - mycelium and conidia - combined with two protocols for DNA extraction using Sodium Dodecyl Sulphate (SDS) and Cetyl Trimethyl Ammonium Bromide as extraction buffers and glass beads for mechanical disruption of cell walls. Our results showed that conidia and SDS buffer was the combination that lead to the best DNA quality and yield, with the lowest variation between samples. This study clearly demonstrates that it is possible to obtain high yield and pure DNA from pigmented conidia without the use of strong cell disrupting procedures and of toxic reagents.
There are numerous methods for DNA extraction from fungi. Some rely on expensive commercial kits and/or equipments, unavailable for many laboratories, or make use of toxic chemicals such as chloroform, phenol and mercaptoethanol. This study clearly demonstrates that it is possible to obtain high yields of pure DNA from pigmented conidia without the use of strong and expensive cell disrupting procedures and of toxic reagents. The method herein described is simultaneously inexpensive and adequate to DNA extraction from several different types of fungi.
本研究的目的是评估一种真菌DNA提取方法，该方法在时间以及昂贵和有毒化学试剂方面投入最少，但能够持续产生高质量的基因组DNA用于PCR。测试了两种类型的真菌生物材料——菌丝体和分生孢子——并结合了两种DNA提取方案，使用十二烷基硫酸钠（SDS）和十六烷基三甲基溴化铵作为提取缓冲液，并使用玻璃珠对细胞壁进行机械破碎。我们的结果表明，分生孢子和SDS缓冲液的组合产生的DNA质量和产量最佳，样品之间的差异最小。这项研究清楚地表明，无需使用强力细胞破碎程序和有毒试剂，就有可能从有颜色的分生孢子中获得高产率和纯净的DNA。
从真菌中提取DNA有许多方法。一些方法依赖于昂贵的商业试剂盒和/或设备，许多实验室无法获得，或者使用氯仿、苯酚和巯基乙醇等有毒化学试剂。这项研究清楚地表明，无需使用强力且昂贵的细胞破碎程序和有毒试剂，就有可能从有颜色的分生孢子中获得高产率的纯净DNA。本文所述方法同时价格低廉，适用于从几种不同类型的真菌中提取DNA。
