Table_2_Taxonomic and Functional Compositions Impacted by the Quality of Metatranscriptomic Assemblies.XLSX

Metatranscriptomics has recently been applied to investigate the active biogeochemical processes and elemental cycles, and in situ responses of microbiomes to environmental stimuli and stress factors. De novo assembly of RNA-Sequencing (RNA-Seq) data can reveal a more detailed description of the metabolic interactions amongst the active microbial communities. However, the quality of the assemblies and the depiction of the metabolic network provided by various de novo assemblers have not yet been thoroughly assessed. In this study, we compared 15 de novo metatranscriptomic assemblies for a fracture fluid sample collected from a borehole located at 1.34 km below land surface in a South African gold mine. These assemblies were constructed from total, non-coding, and coding reads using five de novo transcriptomic assemblers (Trans-ABySS, Trinity, Oases, IDBA-tran, and Rockhopper). They were evaluated based on the number of transcripts, transcript length, range of transcript coverage, continuity, percentage of transcripts with confident annotation assignments, as well as taxonomic and functional diversity patterns. The results showed that these parameters varied considerably among the assemblies, with Trans-ABySS and Trinity generating the best assemblies for non-coding and coding RNA reads, respectively, because the high number of transcripts assembled covered a wide expression range, and captured extensively the taxonomic and metabolic gene diversity, respectively. We concluded that the choice of de novo transcriptomic assemblers impacts substantially the taxonomic and functional compositions. Care should be taken to obtain high-quality assemblies for informing the in situ metabolic landscape.

宏转录组学（Metatranscriptomics）近来已被应用于探究活跃的生物地球化学过程与元素循环，以及微生物组对环境刺激及胁迫因子的原位响应。对RNA测序（RNA-Seq）数据进行从头组装，能够更细致地揭示活跃微生物群落间的代谢互作关系。然而，目前尚未对各类从头组装工具所产出的组装结果质量及代谢网络描绘效果开展全面评估。本研究针对南非某金矿地下1.34千米钻孔采集的裂隙流体样本，对15组宏转录组从头组装结果进行了比较分析。本次组装采用5种从头转录组组装工具（Trans-ABySS、Trinity、Oases、IDBA-tran及Rockhopper），分别基于总读段、非编码读段与编码读段构建而成。评估指标涵盖转录本数量、转录本长度、转录本覆盖度范围、连续性、具备可靠注释的转录本占比，以及分类学与功能多样性模式。研究结果显示，不同组装结果的各项参数差异显著；其中Trans-ABySS与Trinity分别在非编码RNA读段与编码RNA读段的组装中表现最优，前者组装得到的转录本数量众多且覆盖宽泛的表达范围，后者则全面捕获了分类学与代谢基因多样性。本研究证实，从头转录组组装工具的选择会对群落的分类学与功能组成产生显著影响，因此需谨慎选取组装工具以获得高质量结果，从而精准反映原位代谢景观。