This test detects sequence alterations (SNVs, indels) in the coding exons and flanking intronic regions (+/- 10 bp) of POLE (NM_006231.4).
Targeted regions are enriched using hybridization probes (IDT) and sequencing libraries are prepared using the Illumina DNA Prep with Enrichment kit. Paired-end massively parallel sequencing of 150-bp fragments is performed with an Illumina instrument. Sequences are aligned and compared to reference genome GRCh37. Variants are interpreted as per standards and guidelines in the field (PMID: 25741868).
A negative result does not rule out the presence of an alteration that is present below the limits of detection of this assay. The minimum percentage of tumour nuclei required in the submitted sample is 10%; lower percentages may result in false negatives. All samples with less than 10% tumour nuclei will be reported as unsatisfactory. This test does not allow definitive differentiation between germline and somatic variants. Test sensitivity can vary depending on DNA quality, tumour cell percentage, and tumour heterogeneity.
For SNVs and indels, the threshold for variant calling is set at five or more reads, at a minimal sequencing depth of 250 reads, which are the recommended criteria to confidently call variants at an allele frequency of 5% (PMID: 28341590). For DNA sequences, at least 95% of targeted bases must be covered by 250 reads or more to qualify a sample as valid for analysis.
Only clinically relevant variants (i.e. Pathogenic/Likely pathogenic) in the exonuclease domain are reported as suggested (PMID: 31829442). Benign/likely benign variants, variants of uncertain significance with limited evidence for pathogenicity and non-actionable low-risk alleles are not reported.
Turnaround time: 4-6 weeks
Specimens accepted:
- 10 x 10 μm scrolls in 1.5 mL tube
- 10 x 10 μm unstained slides
- A tumour cell content (TCC) of 10% or more is required. This information is mandatory to assess the validity of the test.
Submit your test request using our solid tumour requisition.