demultiplex
nf-core/demultiplex
Demultiplexing pipeline for Illumina sequencing data
Key tools
Run it
nfclaw run demultiplex --input samplesheet.csv --outdir results -profile dockernextflow run pipelines/demultiplex/upstream -profile docker --input samplesheet.csv --outdir resultsnfclaw run demultiplex --demo --outdir results # adds the upstream test profile (-profile test,docker)Inputs
| column | type | required | allowed values | constraints |
|---|---|---|---|---|
id | string | yes | matches ^\S+$ | |
samplesheet | string (file path) | yes | matches ^\S+\.csv$ | |
lane | integer | no | ≥ 1; ≤ 8 | |
flowcell | string (file or directory path) | yes | ||
per_flowcell_manifest | string (file path) | no |
--input must match ^\S+\.csv$.
The samplesheet is a CSV with this header (the columns the schema requires); fill each value per the table above and reference.md (no example value is invented here):
id,samplesheet,flowcellAny of the optional columns above may be appended to the header when your data needs them: lane, per_flowcell_manifest.
From data files. --input may also be the data files — a directory (searched recursively), a quoted glob, or files (--input once per file) — from which nfclaw writes this samplesheet: id from file names; samplesheet from files matching their patterns, by sample name. Set the required columns no file determines with --sheet-set COLUMN=VALUE (a constant or a {sample}/{lane} template): flowcell (string). Preview it with --check; a run records it in <outdir>/provenance/ for replay.
Resources
Resource requests follow the pipeline's configs and selected profiles. nf-core's resource labels often assume a server — a single step can request far more memory than a workstation has, and retry rules can increase requests. If a run fails with Process requirement exceeds available memory (or CPUs), cap every request, and every retry, at what this machine actually has:
nfclaw run demultiplex --input samplesheet.csv --outdir results -profile docker \ --limit-cpus 4 --limit-memory 15.GBnfclaw turns those into Nextflow's process.resourceLimits and passes them as a -c config — the mechanism nf-core prescribes for exactly this (docs). Set them to the machine's real capacity. --limit-time DURATION also caps how long any one task may run: a task that needs longer is stopped, and so is every retry, so set it only as a deliberate ceiling, never as a guess at how long the analysis takes. The generated config is kept in <outdir>/provenance/, so commands.sh replays the run under the same ceiling.
Nextflow engine
This release declares nextflowVersion = '!>=25.10.4'.
To pin this release's declared minimum engine — useful when a newer Nextflow has incompatible config parsing:
nfclaw run demultiplex ... --nxf-ver 25.10.4--nxf-ver is recorded in <outdir>/provenance/, so the replay uses the same engine. See known-issues.
Demo
nfclaw run demultiplex --demo --outdir results # adds the upstream test profile (-profile test,docker)Samplesheet
A CSV with this header (the columns the schema requires). Fill each value per the constraints below; optional columns from the table may be appended.
id,samplesheet,flowcell| Column | Type | Required | Allowed / constraints |
|---|---|---|---|
id | string | required | matches ^\S+$ |
samplesheet | string (file path) | required | matches ^\S+\.csv$ |
lane | integer | optional | ≥ 1; ≤ 8 |
flowcell | string (file or directory path) | required | — |
per_flowcell_manifest | string (file path) | optional | — |
Required parameters
--outdirstring (directory path)The output directory where the results will be saved. You have to use absolute paths to storage on Cloud infrastructure.
--demultiplexerstringDemultiplexer to use.
bases2fastqbcl2fastqbclconvertfqtksgdemuxmkfastqmgikitParameter explorer
44 parameters · 7 groupsEvery parameter from the pinned nextflow_schema.json. nfclaw checks explicit values before launch; complex rules use the pinned pipeline's runtime validation. Search, filter by group, and expand any parameter for its default, allowed values and constraints.
27 parameters · 17 hidden
Checkqc options1
Path to the checkqc config yml file.
Demultiplex options1
Demultiplexer to use.
Downstream Csv options1
Specifies the strandedness of RNA-Seq data for downstream sample sheet generation. This parameter does not affect the demultiplexing process but is used to generate the appropriate field in the nf-core/rnaseq samplesheet
Generic options5
Display the help message.
Display the full detailed help message.
Custom MultiQC yaml file containing HTML including a methods description.
Boolean whether to remove adapter information from Illumina samplesheet. If adapter information is present, the various bcl conversion tools will perform adapter trimming already at the demultiplexing step.
Display hidden parameters in the help message (only works when --help or --help_full are provided).
Input Output options10
Email address for completion summary.
Flowcell ID for single-flowcell runs.
Lane number for single-flowcell runs.
Run directory (or tar.gz) for single-flowcell runs.
Per-flowcell manifest file for fqtk single-flowcell runs.
Path to the flowcell SampleSheet.csv for single-flowcell runs.
Path to comma-separated file containing information about the samples in the experiment.
MultiQC report title. Printed as page header, used for filename if not otherwise specified.
Whether to publish optional outputs such as undertermined files.
The output directory where the results will be saved. You have to use absolute paths to storage on Cloud infrastructure.
Workflow options9
Local JSON file to be passed to samshee module for samplesheet validation
String in JSON format to be passed to samshee module for samplesheet validation
Path to Kraken2 DB to use for screening
Schema name to be passed to samshee module for samplesheet validation
Whether or not to publish raw (untrimmed) FASTQ files alongside trimmed FASTQs. When false, only trimmed FASTQs are published.
Number of reads to subsample for contamination detection.
Comma-separated list of tools to skip (fastp,fastqc,kraken,multiqc,checkqc,falco,md5sum,samshee)
Whether or not to skip trimming
Whether or not illumina samplesheet is v1
Outputs
Results land in --outdir; the files and directory layout depend on the selected workflow, parameters and publication settings. Run metadata is normally placed in <outdir>/pipeline_info/ (execution report, software versions), when those outputs are enabled. The pinned tree includes MultiQC support; a report is produced only when its workflow step runs. nfclaw run also writes <outdir>/provenance/ with the exact params file and the run log, <outdir>/provenance/logs/run.log — the whole launch, whose last line states the outcome (Nextflow's own log is <outdir>/.nextflow.log); unless --no-provenance it adds a run manifest (pinned version, commit and exact command), input/output SHA-256 checksums, and a replayable commands.sh.
The exact output files and directory layout for this release are documented upstream: github.com/nf-core/demultiplex/blob/1.8.0/docs/output.md
Check a run — in the foreground or the background — with nfclaw status <outdir>: success, still running, how it ended (with the error), or stopped without an outcome; exit 0 success, 3 running, 1 otherwise.
Chaining
Run demultiplex as one stage of a chain: nfclaw chain run starts each stage only after the one before it succeeded, and prepares its inputs from that stage's outputs. Every rule is checked against both pinned schemas.
Feeds into
- atacseq — demultiplex writes samplesheet/atacseq_samplesheet.csv (every sample as replicate 1) for its demultiplexed FastQ files; a single-end run's sheet lacks the fastq_2 column, which atacseq's own sheet check requires, so it is added empty.
- methylseq — demultiplex writes samplesheet/methylseq_samplesheet.csv for its demultiplexed FastQ files.
- rnaseq — demultiplex writes samplesheet/rnaseq_samplesheet.csv for its demultiplexed FastQ files (strandedness from its --strandedness, default auto).
- sarek — demultiplex writes samplesheet/sarek_samplesheet.csv with an empty patient; each sample is made its own patient (set it per sample yourself if several samples share one); sarek reads paired-end FastQ only, so every row needs a fastq_2. No status or sex column is written: sarek treats every sample as normal tissue (status 0, sex NA) and runs germline analyses only; a tumour/normal design needs its own sarek samplesheet (a handoff's set gives every row the same value).
- seqinspector — demultiplex writes samplesheet/seqinspector_samplesheet.csv for its demultiplexed FastQ files.
- taxprofiler — demultiplex writes samplesheet/taxprofiler_samplesheet.csv without run_accession or instrument_platform; each sample becomes its own run, sequenced on ILLUMINA (bcl2fastq/bclconvert data — use an inline handoff for other platforms).
nfclaw chain edges demultiplex