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Nextflow Parameters

Complete reference for all pipeline parameters.

Required Parameters

Parameter Description
--input Path to samplesheet CSV
--variants Path to VCF/MAF variants file
--fasta Reference FASTA (with .fai index). Also used for CRAM decoding.

Mode

Parameter Default Description
--mode dna Analysis mode: dna (cfDNA/somatic) or rna (RNA-seq with transcriptome-aware filtering)

Output Options

Parameter Default Description
--outdir results Output directory
--format vcf Output format (vcf or maf)
--suffix '' Suffix for output filenames
--column_prefix '' Prefix for gbcms count columns in MAF output
--preserve_barcode false Keep original Tumor_Sample_Barcode from input MAF

mFSD Options (DNA only)

Parameter Default Description
--mfsd false Enable mFSD analysis — adds 34 mFSD columns to MAF and 7 MFSD_* INFO fields to VCF. See mFSD metrics.
--mfsd_parquet false Write a companion <sample>.fsd.parquet with raw per-variant fragment size arrays. Requires --mfsd.
--mfsd_report false Generate an interactive HTML report with per-variant fragment size distributions. Implies --mfsd and --mfsd_parquet. See mFSD Report.
--mfsd_report_min_alt 3 Minimum ALT fragment count to include a variant in the HTML report.
--mfsd_report_max_variants 20 Maximum variants in the HTML report. Use -1 for no limit.

Quality & Filtering Options

Parameter Default Description
--min_mapq 20 Minimum mapping quality
--min_baseq 20 Minimum base quality
--fragment_qual_threshold 10 Quality margin for fragment consensus — when R1/R2 disagree on a non-INDEL variant, the higher-quality allele wins only if the difference exceeds this. INDEL conflicts with structural CIGAR evidence bypass this threshold.
--context_padding 5 Minimum flanking bases for Phase 3 alignment (auto-increased in repeats)
--adaptive_context true Dynamically increase context padding in tandem repeat regions
--filter_duplicates true Filter duplicate reads
--filter_secondary true Filter secondary alignments
--filter_supplementary true Filter supplementary alignments
--filter_qc_failed true Filter QC failed reads
--filter_improper_pair false Filter improperly paired reads
--filter_indel false Filter reads with indels
--filter_by_sample false Filter multi-sample MAF by Tumor_Sample_Barcode (details)
--show_normalization false Add norm_* columns showing left-aligned coordinates in output
--rescue_mnp false Enable MNP rescue pass — decomposes MNPs into individual SNPs for re-counting when ad=0
--rescue_mnp_threshold 1.0 Maximum disc/len ratio for MNP rescue eligibility (0.0–1.0). 1.0 = all MNPs eligible (C++ compatible). 0.5 = conservative sparse-only mode. 0.0 = disable rescue eligibility (diagnostics still emitted).

UMI & BAQ Options

Parameter Default Description
--umi_tag '' UMI BAM tag for deduplication (e.g., XM, RX). When set, reads sharing the same UMI are grouped as a single observation.
--apply_baq false (DNA) / true (RNA) Apply Base Alignment Quality downgrade. Reduces false positives near indels and splice junctions (CIGAR N). See BAQ Options.

RNA-Specific Options

These parameters are only used when --mode rna is specified.

Parameter Default Description
--rna_editing_db '' Path to REDIportal editing database file (e.g., TABLE1_hg38_v3.txt). Flags ALT sites that overlap known A→I RNA editing positions.
--enforce_strandedness true Enforce dUTP strand-specific library prep. Disable with false for unstranded RNA-seq libraries (--no-strandedness equivalent).

RNA mode defaults

RNA mode uses different PairHMM gap penalties by default (gap_open=5e-3, gap_extend=0.25) to tolerate RT-induced stutter at homopolymers. These can be overridden via the alignment backend parameters below.

Alignment Backend (Advanced)

Parameter Default Description
--alignment_backend pairhmm Phase 3 alignment backend: pairhmm (WFA2 + PairHMM, default) or sw (Smith-Waterman). See CLI Reference.
--llr_threshold 2.3 PairHMM log-likelihood ratio threshold for confident calls
--gap_open_prob 1e-4 PairHMM gap-open probability for non-repeat regions
--gap_extend_prob 0.1 PairHMM gap-extend probability for non-repeat regions
--gap_open_prob_repeat 1e-2 PairHMM gap-open probability for tandem repeat regions
--gap_extend_prob_repeat 0.5 PairHMM gap-extend probability for tandem repeat regions

Merge Options

These parameters control the multi-BAM merge step. Requires a bam_type column in the samplesheet.

Parameter Default Description
--merge_counts false Enable multi-BAM merge — combine per-type MAFs into a single output with type-prefixed count columns. Requires bam_type in samplesheet.
--merge_add_combined true When both duplex and simplex inputs are present, compute additive simplex_duplex_* combined columns (20 columns including strand bias).
--merge_legacy_naming false Use t_{metric}_{type} naming (genotype_variants compatible).

Resource Limits

Parameter Default Description
--max_cpus 16 Maximum CPUs per job
--max_memory 128.GB Maximum memory per job
--max_time 240.h Maximum runtime per job

Performance Benchmarks

Representative metrics from cfDNA duplex BAM samples:

Sample Type BAM Size Variants Runtime CPUs
ctDNA (plasma) 1.3 GB 608 ~25s 4
Plasma control 776 MB 608 ~20s 4

Execution Profiles

Profile Description
docker Local with Docker containers
singularity HPC with Singularity
slurm SLURM cluster with Singularity
local No container (requires local install)

Advanced

task.ext.args — Arbitrary CLI Arguments

Any CLI option not exposed as a Nextflow parameter can be passed via task.ext.args in your config:

process {
    withName: GBCMS_DNA {
        ext.args = '--verbose'
    }
}

See DNA CLI Reference or RNA CLI Reference for all available options.