generate-SQC-BAM

1. Overview
2. How To Run
3. Flow Diagram
4. Pipeline Steps
5. Inputs
6. Outputs
7. Discussions
8. Contributors
9. References

Overview

This pipeline takes BAMs and runs selected Quality Control (QC) steps. Available algorithms are currently SAMtools stats, Picard CollectWgsMetrics and Qualimap bamqc. Generally either Qualimap bamqc or SAMtools stats and Picard CollectWgsMetrics should be run, not both. Qualimap bamqc uses a lot of memory and should not be run within uclahs-cds/metapipeline-DNA. Input can include any combination of tumor and normal BAMs from a single donor. Each will be processed independently. RNA specific QC is not yet implemented but is expected soon.

---

How To Run

1. Update the params section of the .config file (Example config).

2. Update the input YAML (Template YAMLs).

3. See the submission script, here, to submit your pipeline

Requirements

Currently supported Nextflow versions: v23.04.2

---

Flow Diagram

---

Pipeline Steps

Each of the below algorithms, if selected, will run in parallel subject to available resources.

• Note about duplicated reads: SAMtools stats does not ignore reads marked as duplicate by default. The option samtools_remove_duplicates can be set to true to override this. Picard CollectWgsMetrics and Qualimap bamqc do ignore reads marked as duplicate by default.

1. SAMtools stats

samtools stats collects basic statistics from BAM files including read counts, qualities, GC content, insert sizes, read lengths, proper pairing, and duplicated bases.

2. Picard CollectWgsMetrics

picard CollectWgsMetrics collects coverage metrics from WGS BAM files.

3. Qualimap bamqc

qualimap bamqc collects basic statistics and coverage metrics from BAM files. Example output: html pdf. Qualimap bamqc uses a lot of memory and should not be run within uclahs-cds/metapipeline-DNA.

---

Inputs

Input YAML

Example:

---
patient_id: 'patient_id'
dataset_id: 'dataset_id'
input:
  normal:
    - path: /absolute/path/to/normal.bam
      read_length: length
  tumor:
    - path: /absolute/path/to/tumor.bam
      read_length: length

Config

Field	Type	Required	Description
algorithms	list	no	List of tools to be run: ['stats', 'collectwgsmetrics', 'bamqc'], default = ['stats', 'collectwgsmetrics']
reference	path	yes/no	Reference fasta is required only for CollectWgsMetrics
output_dir	path	yes	Not required if blcds_registered_dataset = true
blcds_registered_dataset	boolean	no	Default is false. Only uclahs_cds users should change this. When true, BLCDS folder structure is used
work_dir	path	no	Path of working directory for Nextflow. When included, Nextflow intermediate files and logs will be saved to this directory. With uclahs_cds = true, the default is /scratch and should only be changed for testing/development. Changing this directory to /hot or /tmp can lead to high server latency and potential disk space limitations, respectively.

SAMtools specific configuration

Field	Type	Required	Description
remove_duplicates	boolean	no	Ignore reads marked as duplicate. default = false
samtools_stats_additional_options	string	no	Any additional options recognized by samtools stats

Picard specific configuration

Field	Type	Required	Description
cwm_coverage_cap	integer	no	Cap coverage at this value. Default = 250
cwm_minimum_mapping_quality	integer	no	Ignore reads with mapping quality below this value. Default = 20
cwm_minimum_base_quality	integer	no	Ignore bases with quality below this value. Default = 20
cwm_use_fast_algorithm	boolean	no	If true, fast algorithm is used
cwm_additional_options	string	no	Any additional options recognized by CollectWgsMetrics

Qualimap specific configuration

Field	Type	Required	Description
bamqc_outformat	string	no	Choice of 'pdf' or 'html', default = 'pdf'
bamqc_additional_options	string	no	Any additional options recognized by bamqc

Base resource allocation updaters

To update the base resource (cpus or memory) allocations for processes, use the following structure. The default allocations can be found in the node-specific config files

base_resource_update {
    memory = [
        [['process_name', 'process_name2'], <multiplier for resource>],
        [['process_name3', 'process_name4'], <different multiplier for resource>]
    ]
    cpus = [
        [['process_name', 'process_name2'], <multiplier for resource>],
        [['process_name3', 'process_name4'], <different multiplier for resource>]
    ]
}

Note Resource updates will be applied in the order they're provided so if a process is included twice in the memory list, it will be updated twice in the order it's given.

Examples:

• To double memory of all processes:

base_resource_update {
    memory = [
        [[], 2]
    ]
}

• To double memory for run_CollectWgsMetrics_Picard and triple memory for run_stats_SAMtools and run_bamqc_Qualimap:

base_resource_update {
    memory = [
        ['run_CollectWgsMetrics_Picard', 2],
        [['run_stats_SAMtools', 'run_bamqc_Qualimap'], 3]
    ]
}

• To double CPUs and memory for run_CollectWgsMetrics_Picard and double memory for run_stats_SAMtools:

base_resource_update {
    cpus = [
        ['run_CollectWgsMetrics_Picard', 2]
    ]
    memory = [
        [['run_CollectWgsMetrics_Picard', 'run_stats_SAMtools'], 2]
    ]
}

---

Outputs

Output	Description
{SAMtools-version}_{dataset_id}_{sample_id}_stats.txt	SAMtools stats results
{Picard-version}_{dataset_id}_{sample_id}_wgs-metrics.txt	Picard CollectWgsMetrics results
{Qualimap-version}_{dataset_id}_{sample_id}_stats	Directory of Qualimap results, including, genome_results.txt and either .pdf or .html and supporting directories

---

References

1. SAMtools stats
2. Picard CollectWgsMetrics
3. Qualimap bamqc

---

Discussions

• Issue Tracker to report errors and enhancement ideas.
• Discussions can take place in generate-SQC-BAM Discussions
• generate-SQC-BAM Pull Requests are also open for discussion

---

Contributors

Please see list of Contributors at GitHub.

---

License

Generate-SQC-BAM is licensed under the GNU General Public License version 2. See the file LICENSE for the terms of the GNU GPL license.

Generate-SQC-BAM takes BAM files and generates per sample QC metrics

Copyright (C) 2024 University of California Los Angeles ("Boutros Lab") All rights reserved.

This program is free software; you can redistribute it and/or modify it under the terms of the GNU General Public License as published by the Free Software Foundation; either version 2 of the License, or (at your option) any later version.

This program is distributed in the hope that it will be useful, but WITHOUT ANY WARRANTY; without even the implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the GNU General Public License for more details.