Specimens should be shipped at room temperature in a leak-proof, rigid container with overnight delivery. To discuss minimum acceptable specimens for neonatal patients or for sample types not listed, please contact Allele Diagnostics.
- Peripheral whole blood in EDTA: 2-3 mL
81415 x1, 81416 x2
- Pediatric Exome Test Requisition Form
- Please include clinical notes and pedigree.
Next Generation Sequencing (NGS), including deletion/duplication analysis.
Next Generation Sequencing (NGS) technologies to cover the coding regions of targeted genes plus ~10 bases of non-coding DNA flanking each exon. Patient DNA corresponding to these regions is captured and sequenced. Quality control metrics that are generally achieved: >97% of target bases covered at >20x and mean coverage of target bases >120x. Common benign, likely benign, and low-quality variants are filtered from analysis. Copy number variants (CNVs) are also detected. All reported CNVs are confirmed using another technology such as aCGH, MLPA, or PCR.
Parental samples are sequenced via NGS, and results are compared to the proband results for interpretation purposes. One report is issued for the proband, which includes reference to parental results for variants identified in the proband. For exome testing for proband only, please see test number 8910. For exome using a trio of proband and parents with complete sequencing and reports for each parent, please see test number 8912.
We ask for verification upon test order that patient/guardian consent for all three patients has taken place prior to testing. Reports will consist of five different sections: 1) Variants in genes known to be associated with phenotype, 2) Variants in genes possibly associated with phenotype, 3) Medically actionable variants from the ACMG recommended list of genes (if requested), 4) Variants in genes not associated with phenotype but result in a Mendelian disorder (if requested), 5) Carrier status for variants that are causative for recessive disease (if requested). All differences from the reference sequences (sequence variants) are assigned to one of five interpretation categories (pathogenic, likely pathogenic, variant of uncertain significance, likely benign and benign) per ACMG guidelines. The interpretation of variants is based on current understanding of the genes. These interpretations may change over time as more information about the gene(s) becomes available. Only relevant pathogenic, likely pathogenic, and uncertain variants are reported. A full list of all sequence variants, including likely benign and benign variants, will be provided upon request. Nomenclature for sequence variants comes from Human Genome Variation Society (HGVS). Because no unique report is issued for parents, incidental findings (categories 3-5) are not available for parents with this test. For complete parent reports, see test number 8912.
A small fraction of sequence variants relevant to the proband will not be detected, as some exons cannot be efficiently captured, and some genes cannot be accurately sequenced due to the presence of multiple copies in the genome. Runs of mononucleotide repeats with n >8 in the reference sequence are generally not analyzed. Unless specifically indicated, testing does not include portions of the gene other than coding exons and flanking ~10 bp, such as regulatory domains, deep intronic regions, uncharacterized alternative exons, repeat expansions, epigenetic effects, and mitochondrial genome variants. In most cases, the phase of sequence variants is not determined by this test. The ability to detect minor sequence variants due to somatic mosaicism is limited.
This test detects most deletions and duplications including intragenic CNVs and large cytogenetic events; however, aberrations in a small percentage of regions may not be accurately detected due to sequence paralogy such as pseudogenes or segmental duplications, sequence properties, deletion/duplication size, and inadequate coverage. In general, sensitivity for single, double, or triple exon CNVs is ~70% and for CNVs of 4 exons or larger is >95% but may vary from gene-to-gene based on exon size, depth of coverage, and characteristics of the region. The ability of this assay to detect balanced chromosomal rearrangements, certain types of sex chromosome aneuploidy, exact copy number change within a targeted region, and somatic mosaicism is limited.