Avin Farhat A, Subha Bhassu, Tan Yee-Shin, Braukmann Thomas W A, Vikineswary Sabaratnam, Hebert Paul D N
Mushroom Research Centre (MRC) University of Malaya Kuala Lumpur Malaysia.
Department of Biotechnology Faculty of Science Lincoln University College 47301 Petaling Jaya Malaysia.
Ecol Evol. 2017 Jul 29;7(17):6972-6980. doi: 10.1002/ece3.3049. eCollection 2017 Sep.
DNA barcoding involves the use of one or more short, standardized DNA fragments for the rapid identification of species. A 648-bp segment near the 5' terminus of the mitochondrial cytochrome oxidase subunit I (COI) gene has been adopted as the universal DNA barcode for members of the animal kingdom, but its utility in mushrooms is complicated by the frequent occurrence of large introns. As a consequence, ITS has been adopted as the standard DNA barcode marker for mushrooms despite several shortcomings. This study employed newly designed primers coupled with cDNA analysis to examine COI sequence diversity in six species of and compared these results with those for ITS. The ability of the COI gene to discriminate six species of , the commonly cultivated oyster mushroom, was examined by analysis of cDNA. The amplification success, sequence variation within and among species, and the ability to design effective primers was tested. We compared ITS sequences to their COI cDNA counterparts for all isolates. ITS discriminated between all six species, but some sequence results were uninterpretable, because of length variation among ITS copies. By comparison, a complete COI sequences were recovered from all but three individuals of where only the 5' region was obtained. The COI sequences permitted the resolution of all species when partial data was excluded for . Our results suggest that COI can be a useful barcode marker for mushrooms when cDNA analysis is adopted, permitting identifications in cases where ITS cannot be recovered or where it offers higher resolution when fresh tissue is. The suitability of this approach remains to be confirmed for other mushrooms.
DNA条形码技术涉及使用一个或多个短的、标准化的DNA片段来快速鉴定物种。线粒体细胞色素氧化酶亚基I（COI）基因5'末端附近的一个648碱基对片段已被用作动物界成员的通用DNA条形码，但由于大型内含子频繁出现，其在蘑菇中的实用性变得复杂。因此，尽管存在一些缺点，ITS已被用作蘑菇的标准DNA条形码标记。本研究采用新设计的引物结合cDNA分析来检测6种平菇的COI序列多样性，并将这些结果与ITS的结果进行比较。通过cDNA分析检测了COI基因区分6种常见栽培平菇的能力。测试了扩增成功率、种内和种间的序列变异以及设计有效引物的能力。我们比较了所有分离株的ITS序列与其COI cDNA对应序列。ITS能区分所有6个物种，但由于ITS拷贝之间的长度变异，一些序列结果无法解读。相比之下，除了3个个体外，所有平菇个体都获得了完整的COI序列，这3个个体仅获得了5'区域。当排除平菇的部分数据时，COI序列能够区分所有物种。我们的结果表明，当采用cDNA分析时，COI可以成为蘑菇的有用条形码标记，在无法获得ITS或新鲜组织中ITS提供更高分辨率的情况下允许进行鉴定。这种方法对其他蘑菇的适用性仍有待证实。
