Configuration

Overview

The autoprocessor reads its global settings for the local system from a auto_process.ini file, which it looks for in order in the following locations:

1. The file specified by the AUTO_PROCESS_CONF environment variable (if set)

2. The current directory

3. The config subdirectory of the installation directory

4. The installation directory (for legacy installations only)

Note

In previous versions of the package the configuration file was called settings.ini, and this will be used as a fallback if no auto_process.ini file is found in the locations above.

To create a auto_process.ini file for a new installation, use the command

auto_process.py config --init

To see the current settings, do

auto_process.py config

To change the settings, either use the --set options, for example

auto_process.py config --set bcl2fastq.nprocessors=4

or simply edit the auto_process.ini file by hand using a text editor.

Note

If no auto_process.ini file exists then auto_process will run using the built-in default values.

Note

Many of the configuration options can be over-ridden at run time using command line options for the specific auto_process subcommands.

Basic configuration

Using the basic auto_process Fastq generation requires minimal configuration when running locally; provided that the required BCL conversion software is available on the system (see Software dependencies) it should run without further setup.

Running the QC pipeline requires additional software plus reference data which are described in more detail in Reference data.

Sequencers and platforms

Information about sequencers used at the local site can be stored in sequencer sections of the configuration file.

A sequencer can be defined by adding a new section of the form [sequencer:INSTRUMENT_NAME], where INSTRUMENT_NAME is the ID name for the instrument. Within each section the following data items can then be associated with the sequencer:

platform	Compulsory sets the generic platform name (one of hiseq, hiseq4000, nextseq, miseq, miniseq or iseq)
model	Text describing the sequencer model (e.g. HiSeq 4000)

For example: if the local facility has a HiSeq 4000 instrument with ID SN7001250 then this would be defined in auto_process.ini as follows:

[sequencer:SN7001250]
platform = hiseq4000
model = "HiSeq 4000"

The instrument name can be derived from the name of the directories produced by the sequencer (see Run and Fastq naming conventions).

Note

These sections replace the old sequencers section used to define the sequencer platforms, e.g.

[sequencers]
SN7001250 = hiseq4000

This section is still supported but is now deprecated.

Each platform referenced in the [sequencer:...] sections can optionally be defined in its own [platform:...] section, where platform-specific options for Fastq generation can be set to override those in the [bcl_conversion] section.

The available options are:

bcl_converter	Specify the BCL conversion software to be used when processing data from this platform (see Specifying BCL to Fastq conversion software and options)
nprocessors	Optionally, specify the number of processors to use when performing the BCL to Fastq conversion (deprecated, it is recommended to set this implicitly via the job runners - see Setting number of available CPUs)
no_lane_splitting	Specify whether to merge Fastqs for the same sample across lanes (set to true) or not (set to false)
create_empty_fastqs	Specify whether to create “empty” placeholder Fastqs for samples where demultiplexing failed to assign any reads

For example:

[platform:hiseq4000]
bcl_converter = bcl2fastq>=2.20

Default metadata

The metadata section of the configuration file allows defaults to be specified for metadata items associated with each run.

Currently it is possible to set a default for the source metadata item, which specifies where the data was received from, for example:

[metadata]
default_data_source = "Local sequencing facility"

If no default is set then the values can be updated using the metadata command (see metadata).

Job Runners

Job runners are used within auto_process to tell the pipelines how to execute commands. There are currently two types of runner available:

• SimpleJobRunner runs jobs as a subprocess of the current process, so they run locally (i.e. on the same hardware that the auto_process command was started on)

• GEJobRunner submits jobs to Grid Engine (GE), which enables it to exploit additional resources available on a compute cluster (see Running on a compute cluster)

Job runners can also be configured to specify the number of CPUs available to commands that are executed using them (see Setting number of available CPUs).

By default auto_process is configured to use SimpleJobRunner for all jobs; the default runner is defined in the settings:

[general]
default_runner = SimpleJobRunner

This default can be overridden for specific commands and pipeline stages by explicitly specifying alternative runners in the runners section of the settings file:

Runner name	Used for
barcode_analysis	Running barcode analysis tasks in Fastq generation
bcl2fastq	Running bcl2fastq in Fastq generation
bcl_convert	Running bcl-convert in Fastq generation
cellranger_mkfastq	Running cellranger* mkfastq
cellranger_count	Running cellranger* count
cellranger_multi	Running cellranger multi
stats	Running commands to generate statistics after Fastq generation (e.g. fastq_statistics.py)
rsync	Running commands for transferring data (e.g. copying primary data for Fastq generation, archiving etc)
fastqc	Running FastQC in the QC pipeline
fastq_screen	Running FastqScreen in the QC pipeline
merge_fastqs	Merging Fastq files in Fastq generation
star	Running pipeline tasks which use STAR (e.g. strandedness, alignment etc)
picard	Running Picard in the QC pipeline
qualimap	Running Qualimap in the QC pipeline
rseqc	Running RSeQC geneBody_coverage.py in the QC pipeline
publish_qc	Running jobs for QC publication
icell8	Default runner for commands in the ICELL8 processing pipeline
icell8_contaminant_filter	Running the contaminant filtering in the ICELL8 pipeline
icell8_statistics	Generating statistics for ICELL8 data
icell8_report	Reporting on the ICELL8 pipeline

The following runners are supported but deprecated:

cellranger	Running cellranger in Fastq generation and QC pipelines (used as a fallback for cellranger_* runners)
qc	Running generally computationally intensive QC commands (used as a fallback for fastqc, fastq_screen, star, qualimap and rseqc runners)

Note

It’s recommended to only explicitly configure those runners for which the default runner is not suitable, to avoid a proliferation of unnecessary runner defintions in the configuration file.

Setting number of available CPUs

Job runners allow the number of available CPUs (aka processors or threads) to be specified, and this information is then used when running jobs in the auto_process pipelines.

For SimpleJobRunners the number of CPUs is specified via the nslots argument. For example:

[runners]
qc = SimpleJobRunner(nslots=8)

(Without nslots the number of CPUs implicitly defaults to 1.)

For GEJobRunners the number of available CPUs is inferred from the -pe smp.pe argument (see Running on a compute cluster).

For some commands the number of available CPUs will be taken implicitly from this argument unless explicitly overridden by the following settings:

Section	Setting	Overrides runner
bcl_conversion	nprocessors	bcl2fastq/bcl_convert
fastq_stats	nprocessors	stats
qc	nprocessors	qc
icell8	nprocessors_contaminant_filter	icell8_contaminant_filter
icell8	nprocessors_statistics	icell8_statistics
10xgenomics	cellranger_localcores	cellranger (*)

(*) Used when cellranger is run with --jobmode=local

Running on a compute cluster

The GEJobRunner can be used to make auto_process submit its computationally intensive jobs to a compute cluster rather than on the local host; to switch to using GEJobRunner, set the default runner in the settings:

[general]
default_runner = GEJobRunner

Additional options for Grid Engine submission can be specified by enclosing when defining the runner, for example sending all jobs to a particular queue might use:

default_runner = GEJobRunner(-q ngs.queue)

This default runer can further be overridden for specific commands and pipeline stages by the settings in the runners section of the configuration file (see the previous section Job Runners).

For example: to run bcl2fastq jobs in parallel environment with 8 cores might look like:

[runners]
bcl2fastq = GEJobRunner(-pe smp.pe 8)

Note

If you specify multiple processors for the bcl2fastq runner and are using GEJobRunner then you should ensure that the job runner requests a suitable number of cores when submitting jobs.

Note

When running on a cluster the auto_process driver process should run on the cluster login node; it has a small CPU and memory footprint which should impact minimally on other users of the system.

Managing concurrent jobs and process loads

There are a number of settings available in the [general] section which allow limits to be set on the resources that auto_process will try to consume when running jobs and pipelines:

Setting
max_concurrent_jobs	Maximum number of jobs that auto_process is allowed to run at one time
max_cores	Maximum number of cores that auto_process is allowed to use at one time across all jobs
max_batches	Dynamically sets batch sizes within pipelines so that number of job batches from each task doesn’t exceed this number

For example:

[general]
max_cores = 24

If any of these is set to zero or None then this means that resource is not limited by auto_process.

max_concurrent_jobs and max_batches are useful on shared cluster systems, to avoid submitting large numbers of jobs at one time.

max_cores is useful when running on a local workstation, to avoid exceeding resource limits while ensuring the most efficient use of the available CPUs.

Using environment modules

Environment modules provide a way to dynamically modify the user’s environment. They can be especially useful to provide access to multiple versions of the same software package, and to manage conflicts between packages.

The [modulefiles] section in auto_process.ini allows specific module files to be loaded before a specific step, for example:

[modulefiles]
make_fastqs = apps/bcl2fastq/2.20

These can be defined for the following stages:

• make_fastqs

• run_qc

• publish_qc

• process_icell8

(see Software dependencies for details of what software is required for each of these stages.)

Note

These can be overridden for the make_fastqs and run_qc stages using the --modulefiles option.

Environment modules for Fastq generation

For the make_fastqs stage, additional module files can be specified for individual tasks with the Fastq generation pipeline:

• bcl2fastq

• bcl_convert

• cellranger_mkfastq

• cellranger_atac_mkfastq

• cellranger_arc_mkfastq

If any of these are defined then they will be loaded for the relevant tasks in the Fastq generation pipeline.

Environment modules for the QC pipeline

For the run_qc stage, additional module files can be specified for individual tasks within the QC pipeline:

• fastqc

• fastq_screen

• fastq_strand

• cellranger

• report_qc

If any of these are defined then they will be loaded for the relevant tasks in the QC pipeline.

Note

In older pipeline versions the illumina_qc module file setting was used for the illumina_qc.sh script, which ran both FastQC and FastqScreen. illumina_qc.sh has now been dropped however if the illumina_qc modulefile is still set in the configuration then this will be used as a fallback if fastqc and fastq_screen module files are not set explicitly.

Using conda to resolve pipeline dependencies

For certain pipelines and tasks it is possible to enable the conda package management utility to handle setting up appropriate run-time environments, rather than having to manually install the required dependencies and specify their locations (e.g. using environment modules).

To do this by default, set the enable_conda parameter in the [conda] section, i.e.:

[conda]
enable_conda = true

Note that this requires conda to be installed and available on the user’s PATH at run-time.

By default a temporary directory will be used when creating and reusing conda environments, but this can be overriden by setting the env_dir parameter, e.g.:

[conda]
enable_conda = true
env_dir = $HOME/conda_envs

Specifying BCL to Fastq conversion software and options

The [bcl_conversion] section sets the default settings for BCL to Fastq generation:

bcl_converter	Specify the BCL conversion software to be used when processing data from this platform; see below for more information
nprocessors	Optionally, specify the number of processors to use when performing the BCL to Fastq conversion (deprecated, it is recommended to set this implicitly via the job runners - see Setting number of available CPUs)
no_lane_splitting	Specify whether to merge Fastqs for the same sample across lanes (set to true) or not (set to false)
create_empty_fastqs	Specify whether to create “empty” placeholder Fastqs for samples where demultiplexing failed to assign any reads

Note

This replaces the settings in the old [bcl2fastq] section, which is now deprecated.

The bcl_converter setting can be used to specify both the software package and optionally also a required version; it takes the general form:

bcl_converter = PACKAGE[REQUIREMENT]

Valid package names are:

• bcl2fastq

• bcl-convert

Version requirements are specified by prefacing the version number by one of the operators >, >=, <= and < (== can also be specified explicitly), for example:

bcl_converter = bcl-convert>=3.7

Alternatively a comma-separated list can be provided:

bcl_converter = bcl2fastq>=1.8.3,<2.0

If no version is explicitly specified then the highest available version will be used.

QC pipeline configuration

Several steps in the QC pipeline require reference data to be defined as described in the section run_qc.

Additionally the [qc] section allows other aspects of the QC pipeline operation to be explicitly specified.

Setting size of Fastq read subsets

The default size of the subset of reads used by FastqScreen when generating the screens, generating BAM files and so on can be set using the fastq_subset_size parameter, e.g.:

[qc]
fastq_subset_size = 10000
...

Setting this to zero will force all reads to be used for the appropriate QC stages (note that this can result in extended run time for the QC pipeline, and larger intermediate and final output files).

Note

fastq_subset_size replaces the deprecated legacy fastq_screen_subset parameter (which will however be used as a fallback if subset_size is not present).

Per-lane QC for undetermined Fastqs

By default the QC pipeline will split Fastqs from the undetermined project into separate lanes, in order to generate per-lane metrics.

This is controlled by the split_undetermined_fastqs parameter, which by default is implicitly set as:

[qc]
split_undetermined_fastqs = True
...

To disable the lane splitting, set this parameter to False.

FastqScreen output file naming conventions

By default the QC pipeline creates FastqScreen outputs using the following naming convention:

{FASTQ}_screen_{SCREEN_NAME}.png
{FASTQ}_screen_{SCREEN_NAME}.txt

for example PJB_S1_L001_R1_001_screen_model_organisms.png.

It is possible to revert to the older “legacy” naming convention ({FASTQ}_{SCREEN_NAME}_screen.png etc) by setting the use_legacy_screen_names parameter in the qc section:

[qc]
use_legacy_screen_names = True
...

Data transfer destinations

The transfer_data.py utility can be used to copy Fastqs and other data produced by the auto_process.py pipeline to arbitrary destinations, typically for sharing with end users of the pipeline.

The utility provides a number of command line options to specify a destination and the data that are transferred at runtime. However it is also to define one or more destinations in the configuration file, with appropriate presets for each destination.

A destination can be defined by adding a new section to the config file of the form [destination:NAME], where NAME is the name that will be used to refer to the destination when it is specified in a run of transfer_data.py.

Within each section the following parameters can be set for the destination:

Parameter	Function
directory	Compulsory sets the destination directory to copy files to; can be an arbitrary location of the form [[USER@]HOST:]DIR
subdir	Subdirectory naming scheme
zip_fastqs	Whether to bundle Fastqs into ZIP archives
max_zip_size	Maximum size for each ZIP archive (if Fastqs are bundled)
readme_template	Template file to generate README from
url	Base URL to access copied data at
include_downloader	Whether to include download_fastqs.py
include_qc_report	Whether to include zipped QC reports
hard_links	Whether to hard link to Fastqs rather making copies (for local directories on the same file system as the original Fastqs)

For example:

[destination:webserver]
directory = /mnt/hosted/web
subdir = random_bin
readme_template = README.webserver
url = http://ourdata.com/shared
hard_links = true

See transfer_data.py: copying data for transfer to end users for more information on what these settings do.

Bash tab completion

The auto_process-completion.bash file (installed into the etc/bash_completion.d subdirectory of the installation location) can used to enable tab completion of auto_process.py commands within bash shells.

• For a global installation, copy the file to the system’s /etc/bash_completion.d/ directory, to make it available to all users

• For a local installation, source the file when setting up the environment for the installation (or source it in your ~/.bashrc or similar).