Yes, freebayes-parallel does work, but will only parallelize across a single node. Ideally, and what we're doing here, is allowing Nextflow to handle what jobs are split across -- whether it's a node or multiple nodes.

You provide freebayes-nf with some intervals, like the .fai of your reference and specify a width. Sub-intervals are created of width across your original intervals upon which freebayes will operate.

The resultants VCFs are merged and the final VCF is decomposed and normalized using bcftools.

In practice this looks like:

nextflow run brwnj/freebayes-nf -latest -resume -profile docker \
    --alignments '*.cram' \
    --fasta human.fasta

• --alignments
  • Aligned sequences in .bam and/or .cram format. Indexes (.bai/.crai) must be present.
• --fasta
  • Reference FASTA. Index (.fai) must exist in same directory.

• --outdir
  • Base results directory for output.
  • Default: '/.results'
• --project
  • File prefix for merged and annotated VCF files.
  • Default: 'variants'
• --width
  • The genomic window size per variant calling job.
  • Default: 5000000
• --options
  • Arguments to be passed to freebayes command in addition to those already supplied like --bam, --region, and --fasta-reference.
  • Single quote these when specifying on the command line, e.g. --options '--pooled-discrete'.
  • Default: '--pooled-continuous --pooled-discrete --genotype-qualities --report-genotype-likelihood-max --allele-balance-priors-off --min-alternate-fraction 0.03 --min-repeat-entropy 1 --min-alternate-count 2'
• --intervals
  • Picard-style intervals file to use rather than intervals defined in .fai.
  • Something like Broad's interval lists work here if you want to omit masked regions.
    • See: https://software.broadinstitute.org/gatk/download/bundle
    • And use the wgs_calling_regions file for your genome build.