Comprehensive Transcriptomic Analysis of <i>Cordyceps militaris</i> Cultivated on Germinated Soybeans

The ascomycete fungus Cordyceps militaris infects lepidopteran larvae and pupae and forms characteristic fruiting bodies. Owing to its immune-enhancing effects, the fungus has been used as a medicine. For industrial application, this fungus can be grown on geminated soybeans as an alternative protein source. In our study, we performed a comprehensive transcriptomic analysis to identify core gene sets during C. militaris cultivation on germinated soybeans. RNA-Seq technology was applied to the fungal cultures at seven-time points (2, 4, and 7-day and 2, 3, 5, 7-week old cultures) to investigate the global transcriptomic change. We conducted a time-series analysis using a two-step regression strategy and chose 1460 significant genes and assigned them into five clusters. Characterization of each cluster based on Gene Ontology and Kyoto Encyclopedia of Genes and Genomes databases revealed that transcription profiles changed after two weeks of incubation. Gene mapping of cordycepin biosynthesis and isoflavone modification pathways also confirmed that gene expression in the early stage of GSC cultivation is important for these metabolic pathways. Our transcriptomic analysis and selected genes provided a comprehensive molecular basis for the cultivation of C. militaris on germinated soybeans.蛹虫草（Cordyceps militaris）是一种子囊菌（ascomycete fungus），可侵染鳞翅目幼虫与蛹并形成特征性子实体。因其具有免疫增强活性，该真菌已被作为药物使用。为实现工业化应用，该菌种可在发芽大豆（germinated soybeans）上培养以作为替代蛋白源。本研究针对蛹虫草在发芽大豆上的培养过程开展全面转录组分析，以鉴定其中的核心基因集。我们采用RNA测序（RNA-Seq）技术，对7个时间点（培养2、4、7天以及2、3、5、7周的菌体培养物）进行测序，以解析其全局转录组动态变化。通过两步回归策略开展时间序列分析，最终筛选得到1460个显著差异基因，并将其划分为5个表达聚类簇。基于基因本体（Gene Ontology）与京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes）数据库对各聚类簇进行功能注释与表征后发现，培养两周后菌体的转录谱发生显著改变。对虫草素（cordycepin）生物合成通路与异黄酮修饰通路的基因定位分析同样证实，发芽大豆培养早期的基因表达对上述代谢通路的正常运作至关重要。本研究的转录组分析结果与筛选得到的基因集，为蛹虫草在发芽大豆上的规模化培养提供了全面的分子理论依据。