Steps - Upstream GATK

Alignment

This step uses bwa mem algorithm to align a set of paired-end fastq files to the reference genome. We currently use build hg38/GRCh38, more information here. The output bam file is checked for integrity to ensure there is a properly formatted header and the file is not truncated.

  • CWL: workflow_bwa-mem-to-bam_no_unzip_plus_integrity-check.cwl

Add Read Groups

This step uses AddReadGroups.py (https://github.com/dbmi-bgm/cgap-scripts) to add read groups information to the input bam file based on lanes and flow-cells identifiers. The script extracts read groups information from read names and tags the reads accordingly. Unlike Picard AddOrReplaceReadGroups, which assumes a single read group throughout the file, the script can handle files that contains a mix of multiple lanes and flow-cells. The output bam file is checked for integrity to ensure there is a properly formatted header and the file is not truncated.

  • CWL: workflow_add-readgroups_plus_integrity-check.cwl

Merge BAMs

This step uses Samtools merge to merge multiple bam files when data comes in replicates. If there are no replicates, this step is skipped. The output bam file is checked for integrity to ensure there is a properly formatted header and the file is not truncated.

  • CWL: workflow_merge-bam_plus_integrity-check.cwl

Mark Duplicates

This step uses Picard MarkDuplicates to detect and mark PCR duplicates. It creates a duplicate-marked bam file and a report with duplicate stats. The output bam file is checked for integrity to ensure there is a properly formatted header and the file is not truncated.

  • CWL: workflow_picard-MarkDuplicates_plus_integrity-check.cwl

Sort BAM

This step uses Samtools sort to sort the input bam file by genomic coordinates. The output bam file is checked for integrity to ensure there is a properly formatted header and the file is not truncated.

  • CWL: workflow_sort-bam_plus_integrity-check.cwl

Base Recalibration Report (BQSR)

This step uses GATK BaseRecalibrator to create a base quality score recalibration report for the input bam file.

  • CWL: gatk-BaseRecalibrator.cwl

Apply BQSR and Indexing

This step uses GATK ApplyBQSR to apply the base quality score recalibration report to the input bam file. This step creates a recalibrated bam file and a corresponding index. The output bam file is checked for integrity to ensure there is a properly formatted header and the file is not truncated.

  • CWL: workflow_gatk-ApplyBQSR_plus_integrity-check.cwl