Introduction

zifornd/neopred is a bioinformatics pipeline that performs integrative computational analysis of tumor immunity using bulk RNA-sequencing (RNA-seq) data.

1. Read QC (FastQC)
2. Present QC for raw reads (MultiQC)
3. Alignment (STAR)
4. Sort and index alignments (SAMtools)
5. Quality Control (RSeQC)
6. Gene Quantification (Salmon)
7. Batch Removal (Limma)
8. Duplicate read marking (picard MarkDuplicates)
9. Variant calling (GATK)
10. Variant Annotation
11. Neoantigen detection (arcasHLA)
12. Prioritizing identified neoantigens (pVACseq)

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.

````{=html} First, prepare a samplesheet with your input data that looks as follows:

samplesheet.csv:

csv sample,fastq_1,fastq_2,strandedness,batch,design,sample_group WT_REP1,SRR6357070_1.fastq.gz,SRR6357070_2.fastq.gz,auto,b1,control,WT_REP1_1 WT_REP2,SRR6357072_1.fastq.gz,SRR6357072_2.fastq.gz,reverse,b2,treatment,WT_REP2_1 RAP1_UNINDUCED_REP1,SRR6357073_1.fastq.gz,,reverse,b1,control,SRR6357073_a,RAP1_UNINDUCED_REP1_1

Each row represents a fastq file (single-end) or a pair of fastq files (paired end). Rows with the same sample identifier are considered technical replicates and merged automatically. The strandedness refers to the library preparation and will be automatically inferred if set to auto.The batch refers to a parameter that evaluates if your samples have a batch effect. Design refers to the condition on which to do comparsion. The sample_group refers to condition used in neoanigen for removing duplicates.

--> ````

Now, you can run the pipeline using:

bash nextflow run neopred \ -profile <docker/singularity/podman/shifter/charliecloud/conda/institute> \ --input samplesheet.csv \ --outdir <OUTDIR>

[!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.

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

Pipeline output {#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

zifornd/neopred was originally written by zifo.

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

Contributions and Support

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

Citations

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

This pipeline uses code and infrastructure developed and maintained by the nf-core community, reused here under the MIT license.

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.