nf-core/circrna
Version 1

Workflow Type: Nextflow
Work-in-progress

nf-core/circrna

circRNA quantification, differential expression analysis and miRNA target prediction of RNA-Seq data

Introduction

nf-core/circrna is a best-practice analysis pipeline for the quantification, miRNA target prediction and differential expression analysis of circular RNAs in paired-end RNA sequencing data.

The pipeline is built using Nextflow, a workflow tool to run tasks across multiple compute infrastructures in a very portable manner. It comes with docker containers making installation trivial and results highly reproducible.

Quick Start

  1. Install Nextflow (>=21.04.0)

  2. Install any of Docker, Singularity, Podman, Shifter or Charliecloud for full pipeline reproducibility (please only use Conda as a last resort; see docs)

  3. Download the pipeline and test it on a minimal dataset with a single command:

    nextflow run nf-core/circrna -profile test,
    

    Please check nf-core/configs to see if a custom config file to run nf-core pipelines already exists for your Institute. If so, you can simply use -profile in your command. This will enable either docker or singularity and set the appropriate execution settings for your local compute environment.

  4. Start running your own analysis!

    nextflow run nf-core/circrna -profile  --module 'circrna_discovery, mirna_prediction, differential_expression' --tool 'circexplorer2' --input 'samples.csv' --input_type 'fastq' --phenotype 'phenotype.csv'
    

Refer to usage documentation for exapanded details on running each analysis module.

Pipeline Summary

  1. Input type conversion SamToFastq
  2. Raw read quality control FastQC
  3. Adapter trimming + read filtering BBDUK
  4. circRNA quantification
    1. STAR -> CIRCexplorer2
    2. STAR -> circRNA finder
    3. STAR -> DCC
    4. HISAT2 -> CIRI2 -> BWA -> CIRIquant
    5. Bowtie2 -> find circ
    6. Bowtie -> MapSplice
    7. Segemehl -> Segemehl
  5. circRNA filtering
    1. Filter candidate circRNAs by number of reads spanning back-splice junction
  6. circRNA annotation
    1. Annotate candidates as circRNA, ciRNA or EI-circRNA
    2. Calculate mature spliced length
    3. Export mature spliced length as FASTA file
    4. Annotate parent gene, underlying transcripts.
    5. Export information as customised BED12 file
  7. circRNA count matrix
    1. Combine results of quantification tools to produce counts matrix for downstream statistical analysis
    2. Require circRNAs in matrix to be called by at least n quantification tools (consensus filtering)
  8. miRNA target prediction
    1. miRanda
    2. TargetScan
    3. Filter results, miRNAs must be called by both tools
  9. Differential expression analysis DESeq2
  10. MultiQC report MultiQC

Ouputs given by each step in the pipeline can be viewed in the output documentation

Documentation

The nf-core/circrna pipeline comes with documentation about the pipeline: usage and output.

Credits

nf-core/circrna was originally written by Barry Digby (@BarryDigby) from the National University of Ireland, Galway as a member of Dr. Pilib Ó Broins lab with the financial support of Science Foundation Ireland (Grant number 18/CRT/6214).

Contributions and Support

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

For further information or help, don't hesitate to get in touch on the Slack #circrna channel (you can join with this invite).

Citations

You can cite the nf-core publication as follows:

The nf-core framework for community-curated bioinformatics pipelines.

Philip Ewels, Alexander Peltzer, Sven Fillinger, Harshil Patel, Johannes Alneberg, Andreas Wilm, Maxime Ulysse Garcia, Paolo Di Tommaso & Sven Nahnsen.

Nat Biotechnol. 2020 Feb 13. doi: 10.1038/s41587-020-0439-x.

In addition, references of tools and data used in this pipeline are as follows:

Test data References

Dong Cao (2021). An autoregulation loop in fust-1 for circular RNA regulation in Caenorhabditis elegans. Biorxiv. Available at: https://doi.org/10.1101/2021.03.22.436400.

Version History

dev @ 4c9fe3f (earliest) Created 22nd Feb 2022 at 14:51 by Barry Digby

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Created: 22nd Feb 2022 at 14:51

Last used: 28th Jun 2022 at 16:03

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