milou Pipeline Parameters

This page provides a complete reference for all command-line parameters available in milou. Default values are appropriately showcased.

To modify or adjust any parameters, please edit the

1. conf/params.config,
2. conf/resources.config for resource allocation, and
3. conf/gpu.config for GPU configuration.
4. conf/test_local.config for minimal test data run
5. conf/test_full.config for full test data run
6. conf/benchmark.config for performance tracking and resource usage benchmarking
7. conf/replicate_article.config for replicate article run

User can also adjust the DAG rendering options in conf/dag.config file.

Input/Output Options

Parameter	Description	Type	Default	Required	Hidden?
--sample_sheet	Path to Samplesheet.csv with following headers: sample_id, group, read1, read2, assay_type (optional).	string	null	✔️	❌
--genome_fasta	Path to the reference genome FASTA file.	string	null	✔️	❌
--assay_type	Global chemistry override for clipping offsets ('wgbs', 'emseq', 'twist').	string	'wgbs'	❌	❌
--save_reference	If true, the Bismark/BWA-meth index is saved to a persistent storeDir for reuse.	boolean	false	❌	❌
--bismark_index	Path to a pre-built Bismark index directory.	string	false	❌	❌
--aligned_bams	Start the pipeline from previously aligned BAM files instead of fastQ.	boolean	false	❌	❌
--outdir	Output directory where results will be stored.	string	'results'	✔️	❌
--design_file	Path to a custom design matrix for specialized comparisons.	string	null	❌	✔️

QC & Alignment Parameters

Parameter	Description	Type	Default	Required	Hidden?
--aligner	Aligner to use: 'bismark' or 'bwameth'.	string	'bismark'	❌	❌
--use_parabricks	Set to true to use NVIDIA Parabricks for alignment.	boolean	false	❌	❌
--qualimap_args	Additional arguments to pass to Qualimap.	string	""	❌	✔️
--multiqc_config	multiqc config file. Default is null. --multiqc_config [path/to/config] to use multiqc config file.	string	null	❌	✔️
--multiqc_title	multiqc title. Default is null. --multiqc_title [title] to use multiqc title.	string	null	❌	✔️

Methylation Calling & Extraction

Parameter	Description	Type	Default	Required	Hidden?
--coverage_threshold	Minimum read coverage for analysis AND the threshold for automated Clinical Report Pass/Fail validation.	integer	3	✔️	❌

Differential Methylation Analysis

Parameter	Description	Type	Default	Required	Hidden?
--diff_meth_method	Differential method(s) to use: 'dss', 'edger', 'methylkit', or a comma-separated list.	string	'dss'	✔️	❌
--compare_str	The group comparison string limit (e.g., 'Healthy-Tumor').	string	'all'	✔️	❌
--smoothing	Enable or disable moving-average spline smoothing in DSS. Set to false (--smoothing FALSE) for whole-genome WGBS cohorts (~28M CpGs) to reduce peak RAM from >250 GB to <50 GB.	boolean	true	❌	❌
--skip_diff_meth	Skip the differential methylation stage completely.	boolean	false	❌	❌
--methylkit.assembly	Assembly name for MethylKit context.	string	'hg38'	✔️	❌
--methylkit.diff	Minimum methylation difference percentage for Methylkit.	number	0.05	✔️	❌
--methylkit.qvalue	Maximum Q-value threshold for Methylkit.	number	1	✔️	❌
--methylkit.mc_cores	Number of cores to use for MethylKit.	integer	16	✔️	❌
--methylkit.bed_file	Path to the Twist Target region BED file for MethylKit context.	string	null	❌	❌

Functional Annotation & Enrichment

Parameter	Description	Type	Default	Required	Hidden?
--gtf_file	Path to a GTF annotation file (e.g., Gencode/Ensembl) for gene mapping.	string	null	✔️	❌
--refseq_file	Path to a RefSeq BED file for annotation.	string	null	✔️	❌
--disgenet_db	Path to a sovereign, local DisGeNET TSV release for clinical offline mapping.	string	null	❌	❌
--post_processing	Enable post-processing summaries and visualization.	boolean	true	✔️	❌
--logfc_cutoff	Log2 Fold Change threshold for significance.	number	0.5	✔️	❌
--pvalue_cutoff	P-value threshold for statistical significance.	number	0.05	✔️	❌
--top_n_genes	Number of top genes to include in GO/KEGG enrichment analysis.	integer	100	✔️	❌
--hyper_color	Color for hyper-methylated points in graphs.	string	'red'	✔️	❌
--hypo_color	Color for hypo-methylated points in graphs.	string	'blue'	✔️	❌
--nonsig_color	Color for non-significant points in graphs.	string	'black'	✔️	❌

Clinical Reporting Layer

Parameter	Description	Type	Default	Required	Hidden?
--mode	Pipeline mode: 'research' (standard) or 'clinical' (enforces 2-out-of-3 consensus voting).	string	'research'	❌	❌
--offline	Disables internet access and API queries. Enforced automatically in clinical_offline profile.	boolean	false	❌	❌
--run_clinical_report	Force generation of automated Quarto clinical PDF/HTML report.	boolean	false	❌	❌
--promoter_dist	Distance from TSS (upstream) to define a Promoter region.	integer	2000	❌	❌
--enhancer_dist	Distance from TSS to define an Enhancer/Distal region.	integer	10000	❌	❌

Logging & Resource Management

Parameter	Description	Type	Default	Required	Hidden?
--max_memory	Maximum amount of RAM available for any single task.	string	'128.GB'	❌	✔️
--max_cpus	Maximum number of CPUs available for any single task.	integer	16	❌	✔️
--max_time	Maximum walltime for any single task.	string	'240.h'	❌	✔️
--dag.file	Path to the output DAG file.	string	'workflow_dag.dot'	❌	✔️
--dag.overwrite	Overwrite the output DAG file if it already exists.	boolean	false	❌	✔️
--dag.renderHTML	Render the DAG as HTML.	boolean	true	❌	✔️
--dag.renderFormat	Format to render the DAG.	string	'png'	❌	✔️
--dag.renderOptions	Options to pass to the rendering tool.	string	'-Tpng -Gdpi=300'	❌	✔️
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Generic Pipeline Options

Parameter	Description	Type	Default	Required	Hidden?
-profile	Configuration profile: docker, singularity, gpu, clinical, clinical_offline, benchmark.	string	variable	✔️	❌
-resume	Re-start the pipeline from where it left off.	boolean	false	❌	❌
-w	Custom working directory for intermediate files.	string	'work/'	❌	✔️
--help	Display the pipeline help message.	boolean	false	❌	✔️