Configuration File

The rustqc rna subcommand supports an optional YAML configuration file for advanced settings that go beyond what CLI flags offer. Pass the config file with --config / -c:

rustqc rna sample.bam --gtf genes.gtf -p -c config.yaml -o results/

All sections and fields are optional. Missing fields use their default values. Unknown fields are silently ignored, so config files remain forward-compatible.

The configuration file mirrors the CLI subcommand hierarchy. All RNA-Seq QC settings live under a top-level rna: key, matching the rustqc rna subcommand.

Note

The rna: section controls all tools: dupRadar, featureCounts, all 8 RSeQC tools (including TIN), Qualimap, preseq, and samtools. Each tool has an enabled toggle and tool-specific parameter overrides where applicable. CLI flags take precedence over config file values.

Full example

rna:
  # Library settings
  stranded: unstranded # unstranded, forward, or reverse
  paired: true # Paired-end mode

  # Chromosome name remapping
  chromosome_prefix: "chr"
  chromosome_mapping:
    chrM: "MT"

  # Output settings
  sample_name: "MySample" # Override BAM-derived sample name for output filenames
  flat_output: false # Set to true to skip subdirectories

  # dupRadar output toggles
  dupradar:
    dup_matrix: true
    intercept_slope: true
    density_scatter_plot: true
    boxplot: true
    expression_histogram: true
    multiqc_intercept: true
    multiqc_curve: true

  # featureCounts output toggles
  featurecounts:
    counts_file: true
    summary_file: true
    biotype_summary_file: true
    biotype_counts: true
    biotype_counts_mqc: true
    biotype_rrna_mqc: true
    biotype_attribute: "gene_biotype"

  # RSeQC tool toggles and settings
  bam_stat:
    enabled: true
  infer_experiment:
    enabled: true
    sample_size: 200000
  read_duplication:
    enabled: true
  read_distribution:
    enabled: true
  junction_annotation:
    enabled: true
    min_intron: 50
  junction_saturation:
    enabled: true
    seed: 42
    min_coverage: 1
    percentile_floor: 5
    percentile_ceiling: 100
    percentile_step: 5
  inner_distance:
    enabled: true
    sample_size: 1000000
    lower_bound: -250
    upper_bound: 250
    step: 5

  # TIN (Transcript Integrity Number)
  tin:
    enabled: true
    seed: 12345
    sample_size: 100
    min_coverage: 10

  # Qualimap RNA-Seq QC
  qualimap:
    enabled: true

  # Library complexity (preseq lc_extrap)
  preseq:
    enabled: true
    max_extrap: 10000000000
    step_size: 1000000
    n_bootstraps: 100
    confidence_level: 0.95
    seed: 408
    max_terms: 100
    max_segment_length: 100000000
    defects: false

  # Samtools-compatible outputs
  flagstat:
    enabled: true
  idxstats:
    enabled: true
  samtools_stats:
    enabled: true

Library settings

stranded

Library strandedness for strand-aware read counting:

Value	Meaning
unstranded	Count reads on either strand
forward	Forward stranded (read 1 maps to the transcript strand)
reverse	Reverse stranded (read 2 maps to the transcript strand)

Default: unstranded

This can also be set via the -s / --stranded CLI flag, which takes precedence over the config file value.

paired

Enable paired-end mode. When true, read pairs are counted as a single fragment.

Default: false (single-end mode)

This can also be set via the -p / --paired CLI flag. If either the CLI flag or the config file value is true, paired-end mode is enabled.

rna:
  stranded: reverse
  paired: true

Output settings

sample_name

Override the sample name used for output filenames. By default, the sample name is derived from the BAM file stem (e.g., sample.markdup.sorted.bam produces files named sample.markdup.sorted.*). When set, the provided name is used instead.

Default: derived from BAM filename

This can also be set via the --sample-name CLI flag, which takes precedence over the config file value. Can only be used with a single input file.

rna:
  sample_name: "MySample"

Chromosome name mapping

When the chromosome names in your alignment file differ from those in the GTF, RustQC can remap them automatically. Two mechanisms are available, and they can be combined.

chromosome_prefix

A string prefix to prepend to alignment file chromosome names before matching against GTF names. Applied first, before explicit mapping lookups.

rna:
  # Alignment has "1", "2", "X"; GTF has "chr1", "chr2", "chrX"
  chromosome_prefix: "chr"

chromosome_mapping

An explicit mapping from GTF chromosome names (keys) to alignment file chromosome names (values). Applied after the prefix, so explicit entries can override the prefix for specific chromosomes.

rna:
  # After adding "chr" prefix, override the mitochondrial chromosome
  chromosome_mapping:
    chrM: "MT"

A common use case is GENCODE GTFs (which use chr1, chr2, …) with Ensembl alignments (which use 1, 2, …):

rna:
  chromosome_prefix: "chr"
  chromosome_mapping:
    chrM: "MT"

dupRadar output toggles

The dupradar: section controls which dupRadar output files are generated. All outputs are enabled by default.

rna:
  dupradar:
    dup_matrix: true # Duplication matrix TSV
    intercept_slope: true # Intercept/slope fit results
    density_scatter_plot: true # Density scatter plot (PNG + SVG)
    boxplot: true # Duplication rate boxplot (PNG + SVG)
    expression_histogram: true # Expression histogram (PNG + SVG)
    multiqc_intercept: true # MultiQC intercept/slope file
    multiqc_curve: true # MultiQC fitted curve file

Set any field to false to skip generating that output:

rna:
  dupradar:
    boxplot: false
    expression_histogram: false

featureCounts output toggles

The featurecounts: section controls which featureCounts-compatible output files are generated, plus the biotype attribute setting.

rna:
  featurecounts:
    counts_file: true # featureCounts-compatible counts TSV
    summary_file: true # Gene-level assignment summary
    biotype_summary_file: true # Biotype-level assignment summary
    biotype_counts: true # Biotype counts TSV
    biotype_counts_mqc: true # Biotype counts MultiQC bargraph file
    biotype_rrna_mqc: true # Biotype rRNA percentage MultiQC file
    biotype_attribute: "gene_biotype" # GTF attribute for biotype grouping

biotype_attribute

The GTF attribute name used for biotype grouping. This controls how genes are categorized in the biotype output files.

GTF source	Typical attribute
Ensembl	gene_biotype
GENCODE	gene_type

Default: "gene_biotype"

This can also be set via the --biotype-attribute CLI flag, which takes precedence over the config file value.

RustQC auto-detects the biotype attribute if the specified one is not found in the GTF. If neither gene_biotype nor gene_type is present, a warning is printed and biotype counting is skipped.

RSeQC tool settings

Each of the 8 RSeQC tools has an enabled toggle (default true) and tool-specific parameter overrides. Disabling a tool here prevents it from running even when annotation is provided. CLI flags take precedence over config file values for all parameters.

bam_stat

rna:
  bam_stat:
    enabled: true # Set to false to skip bam_stat

No additional parameters. This tool does not require annotation.

infer_experiment

rna:
  infer_experiment:
    enabled: true
    sample_size: 200000 # Number of reads to sample (default: 200000)

Requires annotation (--gtf). The sample_size can also be set via --infer-experiment-sample-size.

read_duplication

rna:
  read_duplication:
    enabled: true # Set to false to skip read_duplication

No additional parameters. This tool does not require annotation.

CLI shortcut: Use --skip-read-duplication to disable without a config file.

read_distribution

rna:
  read_distribution:
    enabled: true # Set to false to skip read_distribution

No additional parameters. Requires annotation (--gtf).

junction_annotation

rna:
  junction_annotation:
    enabled: true
    min_intron: 50 # Minimum intron length in bases (default: 50)

Requires annotation (--gtf). The min_intron can also be set via --min-intron.

junction_saturation

rna:
  junction_saturation:
    enabled: true
    seed: 42 # Random seed for reproducible sampling (default: 42)
    min_coverage: 1 # Minimum read count to consider a junction (default: 1)
    percentile_floor: 5 # Sampling start percentage (default: 5)
    percentile_ceiling: 100 # Sampling end percentage (default: 100)
    percentile_step: 5 # Sampling step size (default: 5)

Requires annotation (--gtf). These parameters can also be set via CLI flags: --junction-saturation-seed, --junction-saturation-min-coverage, --junction-saturation-percentile-floor, --junction-saturation-percentile-ceiling, --junction-saturation-percentile-step.

inner_distance

rna:
  inner_distance:
    enabled: true
    sample_size: 1000000 # Number of reads to sample (default: 1000000)
    lower_bound: -250 # Histogram lower bound (default: -250)
    upper_bound: 250 # Histogram upper bound (default: 250)
    step: 5 # Histogram bin width (default: 5)

Requires annotation (--gtf). These parameters can also be set via CLI flags: --inner-distance-sample-size, --inner-distance-lower-bound, --inner-distance-upper-bound, --inner-distance-step.

tin

rna:
  tin:
    enabled: true
    seed: 12345 # Random seed for reproducible results (default: none, non-deterministic)
    sample_size: 100 # Equally-spaced positions to sample per transcript (default: 100)
    min_coverage: 10 # Minimum read-start count to compute TIN (default: 10)

Requires annotation (--gtf). The TIN (Transcript Integrity Number) measures transcript integrity via Shannon entropy of read coverage uniformity. The seed can also be set via --tin-seed.

CLI shortcut: Use --skip-tin to disable without a config file.

qualimap

rna:
  qualimap:
    enabled: true # Set to false to skip Qualimap RNA-Seq QC analysis

Requires annotation (--gtf only). Runs the Qualimap RNA-Seq QC analysis: gene body coverage profiling (100 percentile bins, 5’ to 3’), 5’/3’ bias metrics, read origin classification (exonic/intronic/intergenic), strand-specificity estimation, and splice junction motif counting. Produces Qualimap-compatible output files parseable by MultiQC.

preseq

rna:
  preseq:
    enabled: true
    max_extrap: 10000000000 # Maximum extrapolation depth (default: 1e10)
    step_size: 1000000 # Step size between extrapolation points (default: 1e6)
    n_bootstraps: 100 # Bootstrap replicates for confidence intervals (default: 100)
    confidence_level: 0.95 # CI confidence level (default: 0.95)
    seed: 408 # Random seed for reproducibility (default: 408, matching upstream preseq)
    max_terms: 100 # Maximum terms in power series (default: 100)
    max_segment_length: 100000000 # Max merged PE fragment length in bp (default: 1e8)
    defects: false # Use defects model for problematic histograms (default: false)

Only needs BAM fragment info (no annotation required). The max_extrap, step_size, n_bootstraps, seed, and max_segment_length can also be set via CLI flags (--preseq-max-extrap, --preseq-step-size, --preseq-n-bootstraps, --preseq-seed, --preseq-seg-len). Use --skip-preseq to disable entirely.

samtools

flagstat / idxstats / samtools_stats

rna:
  flagstat:
    enabled: true # samtools flagstat-compatible output
  idxstats:
    enabled: true # samtools idxstats-compatible output
  samtools_stats:
    enabled: true # samtools stats compatible output (full format including all histogram sections)

These produce samtools-compatible output files in the samtools/ subdirectory. They share the same BAM statistics accumulator as bam_stat — enabling any of them ensures the statistics are collected.