Clinical exome (WES) – blood relatives

Test covered by the reimbursement:
YES
Clinical expertise code:
208
Test without reimbursement:
YES
Gender:
Woman/Man
Material:
Peripheral blood, Buccal swab …
Turnover time:
6 months
STATIM:
1 month

Material:

Peripheral blood | 1x 3 ml of whole blood in K3 EDTA tube
Storage after examination: week after the report is issued 2 – 8°C
Buccal swab | 2x swab stick for buccal swab collection
Storage after examination: week after the report is issued 2 – 8°C
Isolated DNA from blood | 10–100 ng/μL of isolated DNA from blood in a PCR tube of at least 15 μL.
Storage after examination: stored in a DNA archive without restriction 15°C
Isolated DNA from chorionic villi | 30–100 ng/μL of isolated DNA from chorionic villi in a microtube (Eppendorf type)
Storage after examination: stored in a DNA archive without restriction 15°C
Isolated DNA from amniotic fluid | 30–100 ng/μL of isolated DNA from amniocentesis in a microtube (Eppendorf type)
Storage after examination: stored in a DNA archive without restriction 15°C
Isolated DNA from cordocentesis | 30–100 ng/μL of isolated DNA from cordocentesis in a microtube (Eppendorf type)
Storage after examination: stored in a DNA archive without restriction 15°C
DNA isolated from the product of conception | 50–100 ng/μL in microtube (Eppendorf type)
Storage after examination: stored in a DNA archive without restriction 15°C

Quick test description:

DNA testing of relatives using exome sequencing method to determine causal variants responsible for patient phenotype (duo/trio analysis).

Test details:

The clinical exome is tested by massively parallel sequencing on the Illumina platform (NovaSeq Xplus) from whole-exome sequencing (WES) data using virtual panels. The testing includes an analysis identifying potentially causal variants according to phenotype-dependent prioritisation for given models of inheritance of the family tested. A minimum sequencing depth of 30x is achieved at >98% of target regions. The obtained data are analysed according to our own bioinformatic evaluation, which includes mapping to the GRCh38 genome reference sequence, variant calling, annotation with the current version of Ensembl and filtering of clinically significant variants according to a number of annotation outputs from population, clinical and prediction databases. 

Each of the above steps is supplemented by quality control. Intron variants are assessed for the effect on pre-mRNA splicing in the range of +/- 15bp from the exon boundary. Variants in 5' and 3' UTR regions are reported if there is evidence of their pathogenic impact in clinical databases. Results are reported only in samples that have passed bioinformatic quality control (e.g. phred score quality, depth of sequencing, uniformity of coverage). 

The clinical interpretation of genetic variants detected is performed by experts based on the indication for testing, a review of relevant scientific literature, external databases, our internal clinical genetic database and the manual checking of sequencing data. All available evidence of identified variants was classified according to the MGL Gennet Variant Interpretation Procedure based on IACR**** standards, ACMG guidelines (PMID: 25741868) and specifying procedures (PMID: 28492532, doi: 10.1136/jmedgenet-2020-107248) and HGVS genetic nomenclature. The clinical significance status of the ClinVar database is periodically checked with the release of a new version of Ensembl. For patients whose pathogenicity classification was changed (to class 4/5) with the release of a new version of the database, we issue an updated laboratory report. The laboratory report also incudes incidental findings of pathogenic and probably pathogenic variants (class 4 and 5) in genes according to ACMG (PMID: 27854360) guidelines, in tumour predisposition genes reported by our laboratory as part of the CZECANCA testing, and in genes reported by our laboratory as part of the CarrierTest. The choice of variants for reporting in the patient's clinical report is up to the attending clinical geneticist.

Indication

  • Recurrent pregnancy loss (SAB, UUT for repeated severe phenotype) Trio/Quatro analysis: Mother – Father – Foetus(es) – Offspring without the phenotype 
  • Postnatally for severe phenotype in children and adults Trio/Quatro analysis: Mother – Father – Offspring with the phenotype – Offspring without the phenotype 
  • Postnatally for the occurrence of cancer in family history/at a young age, with a negative result from the CZECANCA panel Duo/Trio/Single: relevant family members with/without the phenotype 
  • Postnatally for the phenotype of an inherited disease requiring complete NGS analysis of multiple genes Duo/Trio/Single: relevant family members with/without the phenotype 

The indication of genetic testing is decided by a clinical geneticist based on an analysis of the patient’s personal and family history.

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