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Kids First Data Resource Center Germline Structural Variant Workflow

The Kids First Data Resource Center (KFDRC) Germline Structural Variant (SV) Caller Workflow is a common workflow language (CWL) implmentation to generate SV calls from an aligned reads BAM or CRAM file. The workflow makes use of Manta and SvABA to call varaiants then annotates these variants using AnnotSV.

Relevant Softwares and Versions

Manta

Manta calls structural variants (SVs) and indels from mapped paired-end sequencing reads. It is optimized for analysis of germline variation in small sets of individuals and somatic variation in tumor/normal sample pairs. Manta discovers, assembles and scores large-scale SVs, medium-sized indels and large insertions within a single efficient workflow. The method is designed for rapid analysis on standard compute hardware: NA12878 at 50x genomic coverage is analyzed in less than 20 minutes on a 20 core server, and most WGS tumor/normal analyses can be completed within 2 hours. Manta combines paired and split-read evidence during SV discovery and scoring to improve accuracy, but does not require split-reads or successful breakpoint assemblies to report a variant in cases where there is strong evidence otherwise. It provides scoring models for germline variants in small sets of diploid samples and somatic variants in matched tumor/normal sample pairs.

SvABA

SvABA is a method for detecting structural variants in sequencing data using genome-wide local assembly. Under the hood, SvABA uses a custom implementation of SGA (String Graph Assembler) by Jared Simpson, and BWA-MEM by Heng Li. Contigs are assembled for every 25kb window (with some small overlap) for every region in the genome. The default is to use only clipped, discordant, unmapped and indel reads, although this can be customized to any set of reads at the command line using VariantBam rules. These contigs are then immediately aligned to the reference with BWA-MEM and parsed to identify variants. Sequencing reads are then realigned to the contigs with BWA-MEM, and variants are scored by their read support.

SvABA is currently configured to provide indel and rearrangement calls (and anything "in between"). It can jointly call any number of BAM/CRAM/SAM files, and has built-in support for case-control experiments (e.g. tumor/normal, or trios or quads). In case/control mode, any number of cases and controls (but min of 1 case) can be input, and will jointly assemble all sequences together. If both a case and control are present, variants are output separately in "somatic" and "germline" VCFs. If only a single BAM/CRAM is present (input with the -t flag), a single SV and a single indel VCF will be emitted.

A BWA-MEM index reference genome must also be supplied with -G.

AnnotSV

AnnotSV is a program designed for annotating and ranking Structural Variations (SV). This tool compiles functionally, regulatory and clinically relevant information and aims at providing annotations useful to i) interpret SV potential pathogenicity and ii) filter out SV potential false positives.

Input Files

  • Universal
    • germline_reads: The germline BAM/CRAM input that has been aligned to a reference genome.
    • indexed_reference_fasta: The reference genome fasta (and associated indicies) to which the germline BAM/CRAM was aligned.
  • AnnotSV
    • annotsv_annotations_dir: These annotations are simply those from the install-human-annotation installation process run during AnnotSV installation (see: https://github.com/lgmgeo/AnnotSV/#quick-installation). Specifically these are the annotations installed with v3.1.1 of the software. Newer or older annotations can be slotted in here as needed.

Output Files

  • Manta
    • manta_svs: Structural Variants called by Manta
    • manta_indels: Small INDELs called by Manta
  • SvABA
    • svaba_svs: Structural Variants called by SvABA
    • svaba_indels: Small INDELs called by SvABA
  • AnnotSV
    • manta_annotated_svs: This file contains all records from the manta_svs that AnnotSV could annotate.
    • svaba_annotated_svs: This file contains all records from the svaba_svs that AnnotSV could annotate.

Basic Info

References