Table 1.
Characteristics of sequencing approaches and NGS pipeline at the time of sample processing within participating centers.
| Site 1 | Site 2 | Site 3 | Site 4 | |
|---|---|---|---|---|
| Characteristics | ||||
| Sequencing approach | WES | WGS* | WES | WGS* |
| Mean depth of coverage | >75× | 30×** | 50× | 30×** |
| Bioinformatic pipeline | GATK HaplotypeCaller (v3.4-46) | In-house developed pipeline26 using bwa-mem for mapping and GATK (v3.8-1-0-gf15c1c3ef) for variant calling | Customized pipeline using bwa-mem 0.7.13-r1126 for mapping, Picard Markduplicates, and GATK HaplotypeCaller (v3.7-0-gcfedb67) for variant calling | In-house developed pipeline using bwa-mem for mapping and GATK/HaplotypeCaller (v. 3.7) for variant calling |
| Variant annotation | In-house pipeline for exome analysis containing variant- and gene-specific information, among which the variant population frequencies from >30,000 in-house exome |
Alissa Interpret 5.1.4 (Agilent Technologies) Alamut Visual v.2.11 (SophiaGenetics) |
Alissa Interpret 5.1.4 (Agilent, Santa Clara, CA, USA) Alamut Visual v.2.11rev |
Alissa Interpret 5.1.4 (Agilent Technologies), Alamut Visual v.2.12 (SophiaGenetics) |
| Databases for variant interpretation | ||||
| Gene-specific mutation database | In-house database (>30,000 exomes) | In-house, VKGL-shared data, HGMD | In-house, VKGL-shared data, HGMD | In-house, VKGL-shared data, HGMD |
| Population databases | gnomAD | GoNL, gnomAD | gnomAD, ESP, and GoNL | HPO, gnomAD, and database of SNPs |
| Aminoacid conservation and Prediction software | Polyphen2, SIFT, Grantham MutationTaster, and PhyloP | Polyphen2, SIFT, and Grantham scores | AlignGVGD, SIFT, MutationTaster, and PolyPhen | PolyPhen, SIFT, Grantham scores, MutationTaster, AlignGVGD, PhyloP, and Annotation Dependent Depletion scores |
| Splice predictors | SpliceSiteFinder-like, MaxEntScan, NNSPLICE, and GeneSplicer | SpliceSiteFinder-like, MaxEntScan, and NNSPLICE, GeneSplicer | SpliceSiteFinder-like, MaxEntScan, and NNSPLICE, GeneSplicer | SpliceSiteFinder-like, MaxEntScan, NNSPLICE, and GeneSplicer |
| Inheritance and phenotype | OMIM | OMIM | OMIM | OMIM and HPO |
*Whole-genome sequencing process was performed by the Hartwig Medical Foundation, Amsterdam, The Netherlands.
**Emergency cases are processed according to other in-house bioinformatic pipelines, and coverage depth may vary.