oist/luscombeu_rrnascan

Introduction

oist/luscombeu_rrnascan is a comprehensive Nextflow pipeline for detecting, extracting, and analyzing ribosomal RNA (rRNA) and internal transcribed spacer (ITS) sequences from genomic assemblies or sequencing reads. The pipeline supports multiple input types: Illumina paired-end reads (assembled via NOVOPlasty), long-read sequencing data (ONT/PacBio), or pre-assembled genome sequences. It searches for SSU (18S), 5.8S, and LSU (28S) rRNA sequences using Infernal's covariance models and automatically extracts the ITS region spanning ITS1, ITS2 and 5.8S.

Pipeline Overview

1. Assembly (if reads provided):

   • Illumina reads → NOVOPlasty (de novo circular assembly)
   • ONT/PacBio reads → NOVOloci (long-read assembly NOT READY)
   • Pre-assembled genomes → Direct input

2. rRNA Detection & Extraction:

   • rRNA discovery (Infernal cmsearch with covariance models)
   • Sequence extraction (EMBOSS seqret)
   • ITS region identification (between rRNA boundaries)

3. Quality Control:

   • Read QC (FastQC)
   • Results summary (MultiQC)

Usage

Note

If you are new to Nextflow and nf-core, please refer to this page on how to set-up Nextflow. Make sure to test your setup with -profile test before running the workflow on actual data.

Prepare Input Samplesheet

Create a CSV file (samplesheet.csv) with your input data:

sample_ID,read_type,read_1,read_2,seed
SAMPLE1,illumina,reads_R1.fastq.gz,reads_R2.fastq.gz,seed.fasta
SAMPLE2,genome,assembly.fasta,,
SAMPLE3,ONT,nanopore_reads.fastq.gz,,

Columns:

• sample_ID: Unique sample identifier
• read_type: One of illumina, ONT, Pacbio, or genome
• read_1: Path to forward reads (FASTQ/FASTA) or assembly (FASTA)
• read_2: Path to reverse reads (FASTQ, optional for Illumina)
• seed: Optional seed sequence for NOVOPlasty (FASTA)

Run Pipeline

nextflow run oist/luscombeu_rrnascan \
   -profile singularity \
   --input samplesheet.csv \
   --outdir results

Key Parameters

# Assembly parameters
--genome_range '6000-12000'    # Expected genome size range for NOVOPlasty
--kmer 33                       # K-mer size for assembly (default: 33)

Warning

Please provide pipeline parameters via the CLI or Nextflow -params-file option. Custom config files including those provided by the -c Nextflow option can be used to provide any configuration except for parameters; see docs.

Output

Results are saved in the --outdir directory with the following structure:

results/
├── assembly/          # NOVOPlasty assemblies (if reads provided)
├── cmsearch/          # rRNA search results (tab-separated)
├── extract/      # Extracted rRNA and ITS sequences (FASTA)
│   ├── *_SSU.fasta    # 18S rRNA sequences
│   ├── *_ITS1.fasta   # ITS1 regions
│   ├── *_5.8S.fasta   # 5.8S rRNA sequences
│   ├── *_ITS2.fasta   # ITS2 regions
│   ├── *_LSU.fasta    # 28S rRNA sequences
│   └── *_rrna.tsv     # Detailed rRNA coordinates
├── fastqc/            # FastQC reports
├── multiqc/           # MultiQC summary
└── pipeline_info/     # Pipeline execution details

Credits

oist/luscombeu_rrnascan was originally written by Johannes Nicolaus Wibisana.

Contributions and Support

If you would like to contribute to this pipeline, please see the contributing guidelines.