Next-Generation Sequencing (NGS) has a wide range of sequencing methods for biological research. Metatranscriptomics sequencing is NGS platform-based approach that can be used to investigate functional activities in a microbial community. This study described a workflow to analyze metatranscriptomics data of a Sequence Read Archive (SRA) file under accession SRA216099. It was transcript data from environmental sample of Ganoderma boninense PER17 derived from GenBank. Some bioinformatics tools were used for this workflow. From this data, 43 411 contigs were assembled using meta-velvet software combined with CAP3 (Contig Assembly Program). Ribosomal contig sequences were annotated using BLASTN against SILVA database while all non-ribosomal contig sequences were annotated using BLASTX against SWISS-PROT and NR databases. Two genera were identified from ribosomal sequences, i.e., Ganoderma, and Ralstonia. Ganoderma had 28 ribosomal and 43 370 non-ribosomal contig sequences. They were annotated as 20 Large Sub-Unit (LSU) and 8 Small Sub-Unit (SSU) for ribosomal sequence and 18 202 as a functional sequence. Ralstonia sequences were also detected in this metatranscriptomics data. It has seven ribosomal contig sequences (6 LSU and 1 SSU) and six contig functional sequences. This workflow revealed that there were 18 202 putative functional sequences of Ganoderma and Ralstonia in this metatranscriptomics data. This workflow had performed well in revealing 18 202 putative functional sequences of Ganoderma and Ralstonia in metatranscriptomics data from environmental samples of fungi.

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