nf-VIF: A Nextflow-based Virus Insertion Finder

Institut Curie - Bioinformatics Core Facility

Introduction

The pipeline is built using Nextflow, a workflow tool to run tasks across multiple computing infrastructures in a very portable manner. It comes with conda / singularity containers making installation trivial and results highly reproducible, and can be run on a single laptop as well as on a cluster.

The current workflow is based on the nf-core best practice. See the nf-core project from details on guidelines.

Pipline summary

This pipeline was designed to process Illumina sequencing data from the HPV capture protocol. Briefly, it allows to detect and genotype the HPV strain(s) available in the samples, and to precisely map the insertion sites on the Human genome.

1. Reads cleaning and qality controls (TrimGalore!, FastQC)
2. HPV Genotyping (Bowtie2)
3. Local alignment on detected HPV strain(s) (Bowtie2)
4. Detection of putative HPV breakpoints using soft-clipped reads
5. Soft-clipped reads alignment on Human genome reference (BLAT)
6. Detection of insertion loci and filtering of the results
7. Presentation of results in a dynamic report (MultiQC)

Quick help

N E X T F L O W  ~  version 19.04.0
Launching `main.nf` [backstabbing_roentgen] - revision: 93bf83bb3b
HPV v1.1.1dev
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Usage:

nextflow run main.nf --reads '*_R{1,2}.fastq.gz' --genome 'hg19'
nextflow run main.nf --samplePlan sample_plan --genome 'hg19'

Mandatory arguments:
  --reads                       Path to input data (must be surrounded with quotes)
  --samplePlan                  Path to sample plan file if '--reads' is not specified
  --genome                      Name of Human genome reference
  -profile                      Configuration profile to use. Can use multiple (comma separated)
                                Available: conda, singularityPath, cluster, test and more.

Options:
  --singleEnd                   Specifies that the input is single end reads

Genome References:              If not specified in the configuration file or you wish to overwrite any of the references.
  --genome                      Name of iGenomes reference
  --bwt2_index                  Path to Bowtie2 index
  --fasta                       Path to Fasta reference (.fasta)
  --blatdb                      Path to BLAT database (.2bit)

HPV References:
  --fastaHpv                    Path to Fasta HPV reference (.fasta)
  --bwt2IndexHpv                Path to Bowtie2 index for all HPV strains
  --bwt2IndexHpvSplit           Path to Bowtie2 index per HPV strain
  --saveReference               Save all references generated during the analysis. Default: False

Advanced options:
  --minMapq                     Minimum reads mapping quality. Default: 0
  --minLen                      Minimum trimmed length sequence to consider. Default: 15
  --minFreqGeno                 Fraction of reads to consider a genotpye. Default: 0.1
  --splitReport                 Generate one report per sample. Default: false
 
Other options:
  --outdir                      The output directory where the results will be saved
  --email                       Set this parameter to your e-mail address to get a summary e-mail with details of the run sent to you when the workflow exits
  -name                         Name for the pipeline run. If not specified, Nextflow will automatically generate a random mnemonic.

Skip options:
  --skipTrimming               Skip trimming step
  --skipFastqc                 Skip quality controls on sequencing reads
  --skipBlat                   Skip Human mapping with Blat
  --skipMultiqc                Skip report

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Available Profiles
  -profile test                Set up the test dataset
  -profile conda               Build a new conda environment before running the pipeline
  -profile toolsPath           Use the paths defined in configuration for each tool
  -profile singularity         Use the Singularity images for each process
  -profile cluster             Run the workflow on the cluster, instead of locally

Quick run

The pipeline can be run on any infrastructure from a list of input files or from a sample plan as follow. Note that by default, all tools are expected to be available from your PATH. See the full documentation for details and containers usage.

Run the pipeline on a test dataset

See the conf/test.conf to set your test dataset.

nextflow run main.nf -profile test

Run the pipeline from a sample plan

nextflow run main.nf --samplePlan MY_SAMPLE_PLAN --genome 'hg19' --outdir MY_OUTPUT_DIR

Run the pipeline on a cluster

echo "nextflow run main.nf --reads '*.R{1,2}.fastq.gz' --genome 'hg19' --outdir MY_OUTPUT_DIR -profile singularity,cluster" | qsub -N illumina-hpv

Documentation

1. Installation
2. Reference genomes
3. Running the pipeline
4. Output and how to interpret the results
5. Troubleshooting

Credits

This pipeline has been set up and written by the sequencing facility, the genetic service and the bioinformatics platform of the Institut Curie (M. Deloger, S. Lameiras, S. Baulande, N. Servant)

Contacts

For any question, bug or suggestion, please, contact the bioinformatics core facility