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首页> 外文期刊>Scientific reports. >Identification of novel and robust internal control genes from Volvariella volvacea that are suitable for RT-qPCR in filamentous fungi
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Identification of novel and robust internal control genes from Volvariella volvacea that are suitable for RT-qPCR in filamentous fungi

机译：的草菇新颖和健全的内部对照基因适合用于RT-qPCR的在丝状真菌中的识别

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摘要

The selection of appropriate internal control genes (ICGs) is a crucial step in the normalization of real-time quantitative PCR (RT-qPCR) data. Housekeeping genes are habitually selected for this purpose, despite accumulating evidence on their instability. We screened for novel, robust ICGs in the mushroom forming fungus Volvariella volvacea. Nine commonly used and five newly selected ICGs were evaluated for expression stability using RT-qPCR data in eight different stages of the life cycle of V. volvacea. Three different algorithms consistently determined that three novel ICGs (SPRYp, Ras and Vps26) exhibited the highest expression stability in V. volvacea. Subsequent analysis of ICGs in twenty-four expression profiles from nine filamentous fungi revealed that Ras was the most stable ICG amongst the Basidiomycetous samples, followed by SPRYp, Vps26 and ACTB. Vps26 was expressed most stably within the analyzed data of Ascomycetes, followed by HH3 and β-TUB. No ICG was universally stable for all fungal species, or for all experimental conditions within a species. Ultimately, the choice of an ICG will depend on a specific set of experiments. This study provides novel, robust ICGs for Basidiomycetes and Ascomycetes. Together with the presented guiding principles, this enables the efficient selection of suitable ICGs for RT-qPCR.
机译：选择适当的内部对照基因（ICG）是实时定量PCR（RT-QPCR）数据标准化的关键步骤。尽管积累了对其不稳定的证据，但习惯性地为此目的选择了家政基因。我们在蘑菇中筛选了蘑菇，形成真菌volvariella volvacea。使用V.Volvacea的生命周期的八个不同阶段的RT-QPCR数据评估九种常用和五种新选择的ICG。三种不同的算法一致地确定了三种新型ICG（SPRyP，RAS和VPS26）在V.Volvacea中表现出最高的表达稳定性。随后分析来自九个丝状真菌的二十四种表达谱的ICG显示RAS是基础肌肉样品中最稳定的ICG，其次是Spryp，VPS26和Actb。 VPS26最稳定地在分析的ascomycetes数据中表达，其次是HH3和β-桶。没有ICG对所有真菌物种普遍稳定，或用于物种内的所有实验条件。最终，ICG的选择将取决于特定的一组实验。本研究为基础霉素和ascometes提供了新颖，强大的ICGS。与所提出的指导原则一起，这使得能够有效选择适合的RT-QPCR ICG。

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