• Filter and trim reads with fastp
• Map to reference genome using BWA mem
• Sort and compress mapped reads with samtools
• Remove optical duplicates using GATK Mark Duplicates
• SV calling with Delly, Lumpy, and Manta
• Merge SV calls with SURVIVOR
• Annotation of results based on intersection with previously identified mouse SVs, genic and exonic regions.

Input data can be raw FASTQ reads, or a previously mapped BAM file. If a BAM file is provided, the workflow begins with the GATK Mark duplicates step.

• --sampleID
  • Default: <STRING>
  • The sample ID for the input data (required).
• --pubdir
  • Default: /<PATH>
  • Description: The directory that the saved outputs will be stored.
• --cacheDir
  • Default: /projects/omics_share/meta/containers
  • Description: This is directory that contains cached Singularity containers. JAX users should not change this parameter.
• -w
  • Default: /<PATH>
  • The directory that all intermediary files and nextflow processes utilize. This directory can become quite large. This should be a location on /fastscratch or other directory with ample storage.
• --keep_intermediate
  • Default: FALSE
  • Controls whether intermediate analysis files are copied to the pubdir. For this pipeline, this includes unsorted and sorted alignment files.

The following parameters include mutually exclusive options for specifying input data, including --fastq1/--fastq2, OR, --bam

• --fastq1
  • Default: /<PATH>
  • The path to a single FASTQ file, or one of a pair of FASTQs for paired-end data.
• --fastq2
  • Default: /<PATH>
  • The path to the second of a pair of FASTQs for paired-end data.
• --bam
  • /<PATH>
  • The path to a BAM input data if alignment has already been performed outside this pipeline.

The following parameters refer to the reference genome.

• --fasta
  • Default: /<PATH>
  • Path to the reference genome in FASTA format.
• --fasta_index
  • Default: /<PATH>
  • Optional paramter to specify index for reference genome. If not provided, pipeline will generate an index.
• --bwa_index
  • Default: /<PATH>
  • Optional paramter to specify BWA indices for alignment. If not provided, pipeline will generate these indices.

The following parameters specify reference data and annotations. The --genome_build paramter controls whether the default versions are GRCm38 or GRCm39:

• --genome_build
  • Default: GRCm38
  • Parameter that controls reference data used for alignment and annotation. GRCm38 is the default value, GRCm39 is an accepted alternate value.
• --exclude_regions
  • /ref_data/ucsc_mm10_gap_chr_sorted.bed
  • BED file that lists the coordinates of centromeres and telomeres to exclude as alignment targets. Note: default path refers to a location within the containers quay.io/jaxcompsci/lumpy-ref_data:0.3.1--refv0.2.0and quay.io/jaxcompsci/delly-ref_data:1.1.6--refv0.2.0, which require this file.
• --sv_ins_ref
  • Default: /ref_data/variants_freeze5_sv_INS_mm39_to_mm10_sorted.bed.gz
  • BED file that lists previously indentified insertion SVs. Note: default path refers to a location within the container quay.io/jaxcompsci/bedtools-sv_refs:2.30.0--refv0.2.0, which requires this file.
• --sv_del_ref
  • Default: /ref_data/variants_freeze5_sv_DEL_mm39_to_mm10_sorted.bed.gz
  • BED file that lists previously indentified deletion SVs. Note: default path refers to a location within the container quay.io/jaxcompsci/bedtools-sv_refs:2.30.0--refv0.2.0, which requires this file.
• --sv_inv_ref
  • Default: /ref_data/variants_freeze5_sv_INV_mm39_to_mm10_sorted.bed.gz
  • BED file that lists previously indentified inversion SVs. Note: default path refers to a location within the container quay.io/jaxcompsci/bedtools-sv_refs:2.30.0--refv0.2.0, which requires this file.
• --reg_ref
  • Default: /ref_data/mus_musculus.GRCm38.Regulatory_Build.regulatory_features.20180516.gff.gz
  • BED file that lists regulatory features. Note: default path refers to a location within the container quay.io/jaxcompsci/bedtools-sv_refs:2.30.0--refv0.2.0, which requires this file.
• --genes_bed
  • Default: /ref_data/Mus_musculus.GRCm38.102.gene_symbol.bed
  • BED file that lists gene symbol IDs and coordinates. Note: default path refers to a location within the container quay.io/jaxcompsci/bedtools-sv_refs:2.30.0--refv0.2.0, which requires this file.
• --exons_bed
  • Default: /ref_data/Mus_musculus.GRCm38.102.exons.bed
  • BED file that lists exons and coordinates. Note: default path refers to a location within the container quay.io/jaxcompsci/bedtools-sv_refs:2.30.0--refv0.2.0, which requires this file.

fastp filtering paramenters:

• --quality_phred
  • Default: 30
  • Quality score threshold.
• --unqualified_perc
  • Default: 30
  • Percent threhold of unqualified bases to pass reads.

SURVIVOR merging parameters:

• --surv_dist
  • Default: 1000
  • Maximum distance between breakpoints for merging SVs.
• --surv_supp
  • Default: 1
  • The number of callers (out of 4) required to support an SV.
• --surv_type
  • Default: 1
  • Boolean (0/1) that requires SVs to be the same type for merging.
• --surv_strand
  • Default: 1
  • Boolean (0/1) that requires SVs to be on the same strand for merging.
• --surv_min
  • Default: 30
  • Minimum length (bp) to output SVs.

Naming Convention	Description
mmrSVD_illumina_report.html	Nextflow autogenerated report
trace.txt	Nextflow trace of processes
${sampleID}/${sampleID}_ILLUMINA_DLM_struct_var.vcf	VCF output combining merged Delly, Lumpy, and Manta calls annotated for overlap with exonic regions
${sampleID}/${sampleID}.survivor_joined_results.csv	Table of SVs annotated with overlaps of previously identified SVs (beck), genes, exons, regulatory regions
${sampleID}/alignments/${sampleID}.md.bam	Analysis-ready alignment of reads
${sampleID}/alignments/${sampleID}.md.bai	Index for analysis-ready alignment of reads
${sampleID}/alignments/${sampleID}.md.metrics	GATK MarkDuplicates log
${sampleID}/alignments/${sampleID}.insert_size.txt	Inferred read insert size
${sampleID}/alignments/mapped_lumpy/${sampleID}_alignBWA_lumpy.bam	Alignment file modified module process LUMPY_PREP
${sampleID}/stats/${sampleID}_fastp_report.html	Filtering and trimming report from fastp
${sampleID}/stats/${sampleID}_fastp_report.json	Filtering and trimming report from fastp
${sampleID}/unmerged_calls/${sampleID}_dellySort.vcf	SV calls from Delly
${sampleID}/unmerged_calls/${sampleID}_lumpySort.vcf	SV calls from Lumpy
${sampleID}/unmerged_calls/${sampleID}_mantaSort.vcf	SV calls from Manta