SV Identification and Assessment
Structural variant (SV) calls that were predicted to affect a gene on the gene list (n = 83) were filtered and assessed according to the quality of the call, rarity of the variant, and biological plausibility of tumor predisposition caused by the variant (Figure S2). We initially filtered SVs called by Canvas and/or Manta to retain those that were predicted to affect at least one exon, occurred at a frequency of less than 1% across all NIHR BioResource Rare Disease samples (n = 9,110), and fulfilled minimum quality criteria (GQ ≥ 30 for Manta, QUAL ≥ 30 for Canvas). Remaining variants were regarded as potentially pathogenic if they affected a gene associated with tumor predisposition in the heterozygous state (unless there was evidence of homozygosity or compound heterozygosity) and fell into one of the following categories: (1) copy-number loss of coding regions of a tumor-suppressor gene, (2) copy-number gain of coding regions of a proto-oncogene, and (3) any SV type with a predicted breakpoint disrupting the gene. Subsequently, these SV calls were reviewed with IGV and excluded if they occurred in a copy-number variation map of the human genome26 (hg19 stringent). The occurrence of tumors associated with disruption of particular genes in individuals harboring suspected SVs was noted in the same manner as for single-nucleotide variants (SNVs) and indels. BAM files corresponding to all suspected deleterious calls were reviewed in IGV. All SVs were confirmed with Sanger sequencing according to standard protocols. Inversions, translocations, and tandem duplications were confirmed by sequencing across breakpoints, whereas deletions were confirmed by fragment size resulting from long-range PCR if sequencing across the breakpoint was not possible. Primer sequences are available on request.