Demultiplex.vsh

Demultiplex.vsh is a workflow for demultiplexing of raw sequencing data. Currently data from Illumina and Element Biosciences sequencers are supported.

Workflow Overview

The workflow executes the following steps:

1. Unpacking the input data (when a TAR archive is provided)
2. Run bclconvert or bases2fastq
3. Run falco and convert Illumina InterOp information to csv
4. Run multiqc to generate a report

Usage

Two variants of the same workflow are provided, depending on the flexibility in the ouput structure required:

• The runner workflow provides a predifined output structure. It requires the minimal amount of parameters to be provided, at the cost of being less flexible. It is located at target/nextflow/runner/main.nf
• The demultiplex workflow (target/nextflow/demultiplex/main.nf) allows for more fine-grained tuning, but required more parameters to be provided.

Test data

We have provided test data at gs://viash-hub-test-data/demultiplex/v3/demultiplex_htrnaseq_meta/SingleCell-RNA_P3_2, but please feel free to bring your own. The URL of the test data can be provided as-is to the workflow, or you can download everything and specify a local path.

Setup

In order to use the workflows in this package, you'll need to do the following:

• Install nextflow
• Install a nextflow compatible executor. This workflow provides a profile for docker.

Setting up SCM

In order to let nextflow use the viash-hub workflows, you need to setup a SCM file. This can be done once by creating $HOME/.nextflow/scm and adding the following:

providers {
   vsh {
    platform = 'gitlab'
    server = "packages.viash-hub.com"
  }
}

Alternatively, a custom location for the SCM file can be specified using the NXF_SCM_FILE environment variable.

You can check if everything is working by getting the --help for a workflow:

nextflow run \
vsh/demultiplex \
-r v0.3.4 \
--help

(Optional) Resource usage tuning

Nextflow's labels can be used to specify the amount of resources a process can use. This workflow uses the following labels for CPU and memory:

• verylowmem, lowmem, midmem, highmem
• verylowcpu, lowcpu, midcpu, highcpu

The defaults for these labels can be found at src/config/labels.config. Nextflow checks that the specified resources for a process do not exceed what is available on the machine and will not start if it does. Create your own config file to tune the labels to your needs, for example:

// Resource labels
withLabel: verylowcpu { cpus = 2 }
withLabel: lowcpu { cpus = 8 }
withLabel: midcpu { cpus = 16 }
withLabel: highcpu { cpus = 16 }

withLabel: verylowmem { memory = 4.GB }
withLabel: lowmem { memory = 8.GB }
withLabel: midmem { memory = 8.GB }
withLabel: highmem { memory = 8.GB }

When starting nextflow using the CLI, you can use -c to provide the file to nextflow and overwrite the defaults.

Example

nextflow run vsh/demultiplex \
-r v0.3.4 \
-main-script target/nextflow/runner/main.nf \
--input "gs://viash-hub-test-data/demultiplex/v3/demultiplex_htrnaseq_meta/SingleCell-RNA_P3_2"  \
--demultiplexer bclconvert \
--publish_dir example_output/ \
-profile docker \
-c labels.config

Acknowledgements

Developed in collaboration with Data Intuitive and Open Analytics.