nf-core/rnavar

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Introduction

nf-core/rnavar is a bioinformatics pipeline for RNA variant calling analysis following GATK4 best practices.

Pipeline summary

1. Merge re-sequenced FastQ files (cat)
2. Read QC (FastQC)
3. (Optionally) Extract UMIs from FASTQ reads (UMI-tools)
4. Align reads to reference genome (STAR)
5. Sort and index alignments (SAMtools)
6. Duplicate read marking (Picard MarkDuplicates)
7. Scatter one interval-list into many interval-files (GATK4 IntervalListTools)
8. Splits reads that contain Ns in their cigar string (GATK4 SplitNCigarReads)
9. Estimate and correct systematic bias using base quality score recalibration (GATK4 BaseRecalibrator, GATK4 ApplyBQSR)
10. Convert a BED file to a Picard Interval List (GATK4 BedToIntervalList)
11. Call SNPs and indels (GATK4 HaplotypeCaller)
12. Merge multiple VCF files into one VCF (GATK4 MergeVCFs)
13. Index the VCF (Tabix)
14. Filter variant calls based on certain criteria (GATK4 VariantFiltration)
15. Annotate variants (snpEff, Ensembl VEP)
16. Present QC for raw read, alignment, gene biotype, sample similarity, and strand-specificity checks (MultiQC, R)

Summary of tools and version used in the pipeline

Usage

First, prepare a samplesheet with your input data that looks as follows:

samplesheet.csv:

sample,fastq_1,fastq_2
CONTROL_REP1,AEG588A1_S1_L002_R1_001.fastq.gz,AEG588A1_S1_L002_R2_001.fastq.gz

Each row represents a fastq file (single-end) or a pair of fastq files (paired end).

Now, you can run the pipeline using:

nextflow run nf-core/rnavar -profile <docker/singularity/podman/shifter/charliecloud/conda/institute> --input samplesheet.csv  --outdir <OUTDIR> --genome GRCh38

For more details and further functionality, please refer to the usage documentation and the parameter documentation.

Pipeline output

To see the results of an example test run with a full size dataset refer to the results tab on the nf-core website pipeline page. For more details about the output files and reports, please refer to the output documentation.

Credits

rnavar was originally written by Praveen Raj and Maxime U Garcia at The Swedish Childhood Tumor Biobank (Barntumörbanken), Karolinska Institutet. Nicolas Vannieuwkerke at CMGG later joined and helped with further development (v 1.1.0 and forward).

Maintenance is now lead by Maxime U Garcia (now at Seqera Labs)

Main developers:

• Maxime U Garcia
• Nicolas Vannieuwkerke

We thank the following people for their extensive assistance in the development of this pipeline:

• Harshil Patel
• Nicolás Schcolnicov
• Ömer An
• Phil Ewels
• Praveen Raj
• Sarah Maman

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 #rnavar channel (you can join with this invite).

Citations

If you use nf-core/rnavar for your analysis, please cite it using the following doi: 10.5281/zenodo.6669636

An extensive list of references for the tools used by the pipeline can be found in the CITATIONS.md file.

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.