Table 2.
Summary of validation for next-generation sequencing TruSight One (TSO) and cardiovascular sub-panels.
| 1-Plex experiment (average of experiments ± SD) | |||||||
|---|---|---|---|---|---|---|---|
| Metricsb | TSO panel (full) | Comprehensive cardiomyopathy (CMP) and reflex | Hypertrophic cardiomyopathy (HCM) | Dilated cardiomyopathy/left ventricular non-compaction (DCM/LVNC) | Arrhythmogenic right ventricular cardiomyopathy (ARVC) | Marfan syndrome/Loeys–Dietz syndrome (MFS/LDS) | Thoracic aortic aneurysms and dissections (TAAD) and reflex |
| Target region mean depth (x) | 301.25 ± 35.56 | 309 ± 37.7 | 277 ± 30.38 | 322 ± 39.71 | 319.75 ± 43.22 | 240.25 ± 28.55 | 292.75 ± 32.84 |
| Fraction of regions target depth ≥15× | 0.97 ± 0.00 | 0.99 ± 0.00 | 1.00 ± 0.00 | 0.99 ± 0.00 | 0.99 ± 0.00 | 0.99 ± 0.00 | 0.99 ± 0.00 |
| Accuracy | 100% | 100% | 100% | 100% | 100% | 100% | 100% |
| Analytical sensitivity | 0.96 ± 0.01 | 0.96 ± 0.02 | 1.00 ± 0.00 | 0.95 ± 0.03 | 0.98 ± 0.04 | 1.00 ± 0.00 | 0.96 ± 0.03 |
| Analytical specificity | 0.89 ± 0.03 | 0.93 ± 0.03 | 0.92 ± 0.04 | 0.93 ± 0.03 | 0.91 ± 0.05 | 1.00 ± 0.00 | 0.99 ± 0.02 |
| False negative (FN) SNP rate | 0.04 ± 0.01 | 0.04 ± 0.02 | 0.00 ± 0.00 | 0.05 ± 0.03 | 0.02 ± 0.04 | 0.00 ± 0.00 | 0.04 ± 0.03 |
| False positive (FP) SNP rate | 0.11 ± 0.03 | 0.07 ± 0.03 | 0.08 ± 0.04 | 0.07 ± 0.03 | 0.09 ± 0.05 | 0.00 ± 0.00 | 0.01 ± 0.02 |
| Overall genotype concordance (OGC) range | 0.81–0.94 | 0.82–0.94 | 0.83–1.00 | 0.81–0.97 | 0.81–1.00 | N/Aa | 0.87–1.00 |
| Non-reference sensitivity (NRS) range | 0.91–0.98 | 0.87–1.00 | 0.94–1.00 | 0.86–1.00 | 0.81–1.00 | N/Aa | 0.89–1.00 |
| Non-reference discrepancy (NRD) range | 0.07–0.20 | 0.06–0.19 | 0.00–0.17 | 0.04–0.20 | 0.00–0.19 | N/Aa | 0.00–0.13 |
| Non-reference genotype concordance (NRGC) range | 0.90–0.97 | 0.87–1.00 | 0.94–1.00 | 0.86–1.00 | 0.81–1.00 | N/Aa | 0.89–1.00 |
| Precision range | 0.85–0.97 | 0.86–0.98 | 0.85–1.00 | 0.87–1.00 | 0.81–1.00 | N/Aa | 0.95–1.00 |
| Number of loci with >15× coverage (length in basepairs)c | Not assessed | 34 (3,329 bp) | 3 (221 bp) | 21 (1,561 bp) | 6 (423 bp) | 1 (100 bp) | 6 (787 bp) |
| Number of SegDups loci (length in basepairs)d | Not assessed | 5 (1,914 bp) 2e (197 bp) |
0 (0 bp) | 2 (208 bp) 0e (0 bp) |
2 (197 bp) | 0 (0 bp) | 6 (1,917 bp) 1e (173 bp) |
|
3-Plex experiment (average of experiments ± SD) | |||||||
| Metricsb | TSO panel | CMP and CMP reflex | HCM | DCM/LVNC | ARVC | MFS/LDS | TAAD and TAAD reflex |
| Target region mean depth (x) | 117.48 ± 11.92 | 118.03 ± 10.42 | 108.41 ± 12.01 | 122.66 ± 10.57 | 124.07 ± 11.58 | 94.48 ± 9.75 | 114.76 ± 11.74 |
| Fraction of regions target depth ≥15× | 0.95 ± 0.01 | 0.98 ± 0.00 | 0.98 ± 0.01 | 0.98 ± 0.00 | 0.97 ± 0.01 | 0.98 ± 0.01 | 0.98 ± 0.00 |
| Accuracy (%) | 100 | 100 | 100 | 100 | 100 | 100 | 100 |
| Analytical sensitivity | 0.96 ± 0.01 | 0.96 ± 0.03 | 0.99 ± 0.02 | 0.94 ± 0.03 | 0.98 ± 0.04 | 0.97 ± 0.09 | 0.97 ± 0.04 |
| Analytical specificity | 0.89 ± 0.03 | 0.93 ± 0.03 | 0.88 ± 0.03 | 0.94 ± 0.03 | 0.91 ± 0.06 | 0.98 ± 0.06 | 0.96 ± 0.04 |
| FN SNP rate | 0.04 ± 0.01 | 0.04 ± 0.03 | 0.01 ± 0.02 | 0.06 ± 0.03 | 0.02 ± 0.04 | 0.03 ± 0.09 | 0.03 ± 0.04 |
| FP SNP rate | 0.11 ± 0.03 | 0.07 ± 0.03 | 0.12 ± 0.08 | 0.06 ± 0.03 | 0.09 ± 0.06 | 0.02 ± 0.06 | 0.04 ± 0.04 |
| OGC range | 0.81–0.94 | 0.82–0.94 | 0.83–1.00 | 0.81–0.97 | 0.81–1.00 | N/Aa | 0.87–1.00 |
| NRS range | 0.91–0.98 | 0.87–1.00 | 0.94–1.00 | 0.86–1.00 | 0.81–1.00 | N/Aa | 0.89–1.00 |
| NRD range | 0.07–0.20 | 0.06–0.19 | 0.00–0.17 | 0.04–0.20 | 0.00–0.19 | N/Aa | 0.00–0.13 |
| NRGC range | 0.90–0.97 | 0.87–1.00 | 0.94–1.00 | 0.86–1.00 | 0.81–1.00 | N/Aa | 0.89–1.00 |
| Precision range | 0.85–0.97 | 0.86–0.98 | 0.85–1.00 | 0.87–1.00 | 0.81–1.00 | N/Aa | 0.95–1.00 |
aQuantitative measurements OGC, NRS, NRD, NRGC, and precision were not computed for the MFS/LDS panel since only three variants were available for these analyses, and there was not enough power for the performance of the quantitative measurements above. Qualitative measurements of quality control for library preparation, sequencing performance, coverage, and SNP performance indicated that the variability among the runs listed above were within acceptable range.
bSamples MotherLP and ProbandJP were not used in the calculations of FN SNP rate, FP SNP rate, analytic sensitivity, analytic specificity, OGC, NRS, NRD, NRGC, and precision range. Only GenReg samples were used in accuracy calculations.
cNumber of loci (with corresponding length in basepairs in parenthesis) that presented with lower than 15× coverage after 1-plex runs.
dNumber of loci (with length in basepairs in parenthesis) that mapped to a region of known segmental duplications (SegDups).
eNumber of loci (with corresponding number of basepairs in parenthesis) with known SegDup that were successfully validated to be unambiguously amplified and sequenced out of the total regions and corresponding basepairs listed.