Abstract
BackgroundThe debate regarding the relative merits of whole genome sequencing (WGS) versus exome sequencing (ES) centers around comparative cost, average depth of coverage for each interrogated base, and their relative efficiency in the identification of medically actionable variants from the myriad of variants identified by each approach. Nevertheless, few genomes have been subjected to both WGS and ES, using multiple next generation sequencing platforms. In addition, no personal genome has been so extensively analyzed using DNA derived from peripheral blood as opposed to DNA from transformed cell lines that may either accumulate mutations during propagation or clonally expand mosaic variants during cell transformation and propagation.MethodsWe investigated a genome that was studied previously by SOLiD chemistry using both ES and WGS, and now perform six independent ES assays (Illumina GAII (x2), Illumina HiSeq (x2), Life Technologies' Personal Genome Machine (PGM) and Proton), and one additional WGS (Illumina HiSeq).ResultsWe compared the variants identified by the different methods and provide insights into the differences among variants identified between ES runs in the same technology platform and among different sequencing technologies. We resolved the true genotypes of medically actionable variants identified in the proband through orthogonal experimental approaches. Furthermore, ES identified an additional SH3TC2 variant (p.M1?) that likely contributes to the phenotype in the proband.ConclusionsES identified additional medically actionable variant calls and helped resolve ambiguous single nucleotide variants (SNV) documenting the power of increased depth of coverage of the captured targeted regions. Comparative analyses of WGS and ES reveal that pseudogenes and segmental duplications may explain some instances of apparent disease mutations in unaffected individuals.
Highlights
The debate regarding the relative merits of whole genome sequencing (WGS) versus exome sequencing (ES) centers around comparative cost, average depth of coverage for each interrogated base, and their relative efficiency in the identification of medically actionable variants from the myriad of variants identified by each approach
A median of 91,573 SNPs were recovered by the different ES experiments, not all of the SNPs recovered per exome run are coding exonic variants
This may be true because most WGS studies are typically implemented at average depths of coverage well below any current ES approach, due to the theoretical assumption and observations that an approximately 30x average depth of coverage across the whole genome would provide enough read depth to identify >99% of the homozygous variants and approximately 98% of the heterozygous variants; that is, NOT identifying 1% to 2% of variants in the coding regions
Summary
The debate regarding the relative merits of whole genome sequencing (WGS) versus exome sequencing (ES) centers around comparative cost, average depth of coverage for each interrogated base, and their relative efficiency in the identification of medically actionable variants from the myriad of variants identified by each approach. We previously applied WGS using massively parallel next-generation sequencing (NGS) methods to a subject with molecularly undefined, autosomal recessive, Charcot-Marie-Tooth (CMT) neuropathy and identified apparently causative compound heterozygous SH3TC2 variants: nonsense p.R954X The two alleles satisfied multiple criteria for disease causation: both were validated with alternate technologies; the nonsense p.R954X allele had been previously reported in unrelated CMT families and corroborated with functional studies; each segregated faithfully in the pedigree and no other lesion was discovered in the locus despite adequate coverage of all coding bases with NGS reads. Each allele faithfully independently segregated with additional different mild phenotypes observed in other family members susceptibility to carpal tunnel syndrome (presumably due to loss-of-function nonsense mutation) and an electrophysiologically identified subclinical axonopathy (missense allele)
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