nf-core pipeline · denovo-assembly, rna-seq, transcriptome
nf-core/denovotranscript
A de novo transcriptome assembly pipeline for paired-end bulk RNA-seq data without a reference genome.
What nf-core/denovotranscript is
nf-core/denovotranscript is a bioinformatics pipeline designed for de novo transcriptome assembly from paired-end short-read bulk RNA sequencing data. Developed by Avani Bhojwani and Timothy Little as part of the nf-core initiative, it provides an automated solution for reconstructing transcripts when a reference genome is unavailable.
The pipeline processes paired-end FASTQ files through read quality control, trimming, contaminant removal, de novo assembly, redundancy reduction, assembly quality assessment, and transcript abundance quantification. It produces assembled transcript sequences, abundance matrices, and visual summary reports across all processing steps.
What the pipeline does
Input sequencing reads provided via a CSV samplesheet undergo initial quality control with FastQC, followed by adapter and quality trimming using fastp and a second round of FastQC. Users can optionally remove ribosomal RNA or mitochondrial sequences using SortMeRNA.
Transcriptome assembly is performed using any combination of Trinity (with normalised or non-normalised reads) and rnaSPAdes (with soft, medium, or hard filtering options; Trinity with normalised reads and rnaSPAdes with medium filtering are enabled by default). Assembled transcripts undergo redundancy reduction using EvidentialGene tr2aacds, and a transcript-to-gene mapping is generated using gawk.
Assembly quality and completeness are evaluated using BUSCO, rnaQUAST, and TransRate. Finally, transcript abundance is quantified through pseudo-alignment using Salmon, and all read QC, BUSCO, and Salmon metrics are compiled into an interactive MultiQC HTML report.
Key facts
- Pipeline framework
- Nextflow (nf-core)
- Latest release
- 1.2.1 (2025-08-28)
- Input format
- Paired-end FASTQ files via a CSV samplesheet
- Output formats
- FASTA (assembled transcriptome), TSV (transcript abundance), HTML (MultiQC report)
- Assemblers supported
- Trinity, rnaSPAdes
- Redundancy reduction tool
- EvidentialGene tr2aacds
- Quantification tool
- Salmon
What you can use it for
- Non-model organism transcriptome assembly
- Reconstruct full-length transcript sets from bulk RNA-seq when no reference genome is available.
- Multi-assembler consensus generation
- Combine assemblies generated from Trinity and rnaSPAdes, followed by EvidentialGene redundancy reduction to produce a refined final transcriptome.
- Transcript expression quantification
- Pseudo-align trimmed RNA-seq reads back to the newly assembled transcriptome using Salmon to generate transcript abundance estimates.
Run it here
The research agent runs nf-core/denovotranscript with Nextflow on its own machine — up to 32 cores, 256 GiB of memory and 24 hours per run, which covers the test profile and real single-sample runs. It builds the samplesheet, checks it against the pipeline's schema and reads the results back. When a run is too big for one machine, it gives you the exact command for your own cluster instead.
Strengths and limits
Good at
- Supports combining assemblies from multiple assemblers (Trinity and rnaSPAdes) into a single non-redundant set using EvidentialGene tr2aacds.
- Incorporates extensive multi-tool quality validation using BUSCO, rnaQUAST, and TransRate alongside standard read-level QC.
- Outputs an aggregated MultiQC report summarising raw reads, trimmed reads, assembly completeness, and Salmon quantification metrics.
Watch out for
- Supports only paired-end short reads from bulk RNA-seq.
- TransRate assembly evaluation is not executed when running under conda or mamba profiles.
Questions
Can I run nf-core/denovotranscript without my own cluster?
The research agent runs nf-core/denovotranscript with Nextflow on its own machine — up to 32 cores, 256 GiB of memory and 24 hours per run, which covers the test profile and real single-sample runs. It builds the samplesheet, checks it against the pipeline's schema and reads the results back. When a run is too big for one machine, it gives you the exact command for your own cluster instead.
What licence is nf-core/denovotranscript under?
MIT, like every nf-core pipeline.
What input data does the pipeline require?
It requires a CSV samplesheet detailing sample names alongside paths to paired-end FASTQ files (fastq_1 and fastq_2).
Which assemblers are used by default?
By default, the pipeline runs Trinity with normalised reads and rnaSPAdes with medium filtering.
Can ribosomal RNA or mitochondrial contamination be removed?
Yes, SortMeRNA can optionally be run prior to assembly to filter out rRNA or mitochondrial DNA reads.
Why did TransRate skip execution during my run?
TransRate is omitted automatically if the workflow is run using the conda or mamba profiles.
How to cite it
10.5281/zenodo.13324371
Written from nf-core/denovotranscript's own README. Statements citing a figure that does not appear there are removed before publication. Licence and running details come from our registry.