Next Generation Sequencing Service at SDGS

Transcription

1 Next Generation Sequencing Service at SDGS Testing Workflow Library Preparation SureSelect TruSight Shearing of genomic DNA using the Covaris E220 sonicator. End repair, A tailing and ligation of adaptors using SureSelectXT library system (Agilent Technologies). Enrichment by SureSelect target enrichment (Agilent Technologies) using custom in house designed probes. Samples have barcode tags added following target enrichment. Sequencing on the Illumina HiSeq using the HiSeq Rapid SBS Kit v2 performing 2 x 108 bp paired end reads. Genomic DNA fragmented, barcode tagged and captured using the TruSight TM Rapid Capture (Illumina) protocol. Sequencing on the Illumina MiSeq using the MiSeq Reagent Kit v2 performing 2 x 150 bp end paired reads. Data Analysis Based on the open source Best Practices workflow by the Broad Institute (for additional information, see BWA alignment of reads to human genome build hg19. Generation of depth of coverage reports. Checked using Alamut Visual version 2.7 (Rev 2) (Interactive Biosoftware). A minimum threshold of 30-fold read depth is set for exonic sequences and intronic sequences upto and including 5 bp from exon. A minimum threshold of 18-fold read depth is set for intronic sequences from 6 bp to 25 bp from exon. Identification of variants using HaplotypeCaller. Annotation from dbsnp and COSMIC (currently dbsnp138 and COSMIC v67 but updated with new releases) Page 1 of 22

2 Variants filtered against in-house polymorphism lists and Best Practice Guidelines for the evaluation of pathogenicity and the reporting of sequence variants in clinical molecular genetics (Association for Clinical Genetic Science). Post analysis Confirmation of clinically significant sequence variants by Sanger sequencing as necessary (see below). Filling of gaps with low depth of coverage by Sanger sequencing as necessary. Gaps that fall within genes that have an individual whole gene sequencing service listed on the UKGTN service website as filled. The locations of all gaps that are not filled by Sanger sequencing are clearly indicated on the report. Creation of a diagnostic report combining clonal and Sanger sequence data that includes clinical interpretation of all the variants identified that are clearly pathogenic, likely to be pathogenic or have uncertain clinical significance. Confirmation of variants detected by NGS A risk assessment was carried out in June 2016 regarding the necessity for confirming variants identified by NGS using orthogonal technology (typically Sanger sequencing). The risk assessment was based on best practice guidelines, analysis of the quality of NGS data produced and the robustness of the tube transfer checking process within the laboratory. The conclusion was: - SDGS will no longer performs confirmation of single nucleotide substitution variants rated as class 3, 4 and 5, with a QUAL score of 3000 (and depth of coverage of 30x [ 18x for -25_-6 and +6_+25]), when detected by SureSelect and TruSight NGS workflows run on the MiSeq and HiSeq2500 platforms. Exclusions to this include: - Indel variants - these are confirmed by Sanger sequencing, regardless of their QUAL score. AmpliSeq/PGM workflow all variants are confirmed by Sanger sequencing. Rare genes with highly homologous pseudogenes Somatic cancer panels Page 2 of 22

3 Inborn Errors of Metabolism (SureSelect Design ID: ) Glycogen Storage Diseases (GSDs) Condition Gene Subpanel(s) Primary transcript Glycogen storage disease III AGL Liver/Muscle/Heart NM_ Glycogen storage disease XII ALDOA Muscle NM_ Glycogen storage disease XIII ENO3 Muscle NM_ Epilepsy, progressive myoclonic 2A (Lafora) EPM2A Generalised NM_ Fructose-1,6-bisphosphatase deficiency FBP1 Liver NM_ N/A - no mutations reported in gene to date FBP2 Muscle NM_ Glycogen storage disease Ia G6PC Liver NM_ Glycogen storage disease II GAA Generalised/Muscle NM_ Glycogen storage disease IV / Adult polyglucosan body disease GBE1 Liver/Muscle/Heart NM_ Glycogen storage disease XV GYG1 Muscle/Heart NM_ N/A - no mutations reported in gene to date GYG2 Liver NM_ Glycogen storage disease 0, muscle GYS1 Muscle/Heart NM_ Glycogen storage disease, type 0 GYS2 Liver NM_ Danon disease LAMP2 Generalised NM_ Glycogen storage disease XI LDHA Muscle NM_ Epilepsy, progressive myoclonic 2B (Lafora) NHLRC1 Generalised NM_ Hemolytic anaemia due to phosphofructokinase deficiency PFKL Liver NM_ Glycogen storage disease VII PFKM Muscle NM_ Glycogen storage disease X PGAM2 Muscle NM_ Phosphoglycerate kinase 1 deficiency PGK1 Muscle NM_ GSD XIV, Phosphoglucomutase 1 deficiency PGM1 Muscle NM_ GSD Ixd; muscle phosphorylase kinase deficiency PHKA1 Muscle NM_ Glycogen storage disease IXa1; GSD9A1 PHKA2 Liver NM_ Glycogen storage disease IXb; GSD9B PHKB Liver NM_ N/A - no mutatiopns reported in gene to date PHKG1 Muscle NM_ Glycogen storage disease IXc PHKG2 Liver NM_ Glycogen storage disease of heart, lethal congenital PRKAG2 Heart NM_ Glycogen storage disease VI PYGL Liver NM_ Glycogen storage disease, type V; McArdle disease PYGM Muscle NM_ RBCK1 deficiency RBCK1 Muscle/Heart NM_ Fanconi-Bickel syndrome SLC2A2 Liver NM_ Glycogen storage disease Ib / Ic SLC37A4 Liver NM_ genes Page 3 of 22

4 Fatty Acid Metabolism Disorders Condition Gene Subpanel(s) Primary transcript ACAD9 deficiency (complex I deficiency) ACAD9 FAO NM_ MCAD deficiency ACADM FAO NM_ SCAD deficiency ACADS SCAD NM_ VLCAD deficiency ACADVL FAO NM_ Beta-ketothiolase deficiency ACAT1 Ketolysis NM_ CPTI deficiency CPT1A FAO NM_ CPTI deficiency (muscle) CPT1B FAO NM_ CPTII deficiency CPT2 FAO NM_ MADD ETFA FAO NM_ MADD ETFB FAO NM_ MADD (riboflavin-responsive) ETFDH FAO NM_ HADH (SCHAD) deficiency HADH FAO NM_ Mitochondrial TFP deficiency HADHA FAO NM_ Mitochondrial TFP deficiency HADHB FAO NM_ Mitochondrial HMG-CoA lyase deficiency HMGCL FAO NM_ Mitochondrial HMG-CoA synthase deficiency HMGCS2 FAO NM_ SCOT deficiency OXCT1 Ketolysis NM_ Systemic primary carnitine deficiency (carnitine transporter) SLC22A5 FAO NM_ CACT deficiency SLC25A20 FAO NM_ Riboflavin deficiency (RFVT1) SLC52A1 FAO NM_ Brown-Vialetto-van Laere syndrome (RFVT2) SLC52A2 FAO NM_ Brown-Vialetto-van Laere syndrome (RFVT3) SLC52A3 FAO NM_ FAO - Fatty acid oxidaton/ketogenesis Ketolysis - Ketolysis 22 genes SCAD - Suspected SCAD (by specific request secondary to/combined with FAO panel) Page 4 of 22

5 Peroxisomal Disorders Condition Gene Primary transcript X-linked adrenoleukodystrophy ABCD1 NM_ Peroxisomal Acyl-CoA oxidase deficiency ACOX1 NM_ Rhizomelic chondrodysplasia punctata type 3 (RCDP3) AGPS NM_ Type I primary hyperoxaluria (HP1) AGXT NM_ Alpha-methylacyl-CoA racemase (AMACR) deficiency AMACR NM_ Encephalopathy, lethal, due to defective mitochondrial peroxisomal fission DNM1L NM_ Rhizomelic chondrodysplasia punctata type 2 (RCDP2) (DHAPAT deficiency) GNPAT NM_ D-Bifunctional protein deficiency HSD17B4 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX1 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX2 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX3 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX5 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX6 NM_ Rhizomelic chondrodysplasia punctata type 1 (RCDP1) PEX7 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX10 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX11B NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX12 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX13 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX14 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX16 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX19 NM_ Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum PEX26 NM_ Adult Refsum disease PHYH NM_ Leukoencephalopathy with dystonia and motor neuropathy SCP2 NM_ genes Page 5 of 22

6 Hyperammonaemia / Urea Cycle Disorders Condition Gene Subpanel(s) Primary transcript Arginase deficiency (hyperargininemia) ARG1 UCD/Hyperamm NM_ Argininosuccinic aciduria ASL UCD/Hyperamm NM_ Citrullinemia type I ASS1 UCD/Hyperamm NM_ Carbamoylphosphate synthetase I (CPS1) deficiency CPS1 CPS_NAGS/UCD/ Hyperamm NM_ Hyperinsulinism-hyperammonemia syndrome GLUD1 Hyperamm NM_ MMA (mutase) MUT Hyperamm NM_ N-acetyl glutamate synthetase (NAGS) deficiency NAGS CPS_NAGS/UCD/ Hyperamm NM_ Ornithine aminotransferase (OAT) deficiency OAT Hyperamm NM_ Ornithine transcarbamylase (OTC) deficiency OTC UCD/Hyperamm NM_ PA PCCA Hyperamm NM_ PA PCCB Hyperamm NM_ Citrullinaemia type 2 (citrin deficiency) SLC25A13 UCD/Hyperamm NM_ HHH syndrome SLC25A15 UCD/Hyperamm NM_ Lysinuric protein intolerance SLC7A7 Hyperamm NM_ genes CPS_NAGS - CPS1 and NAGS only UCD - Urea cycle disorders Hyperamm - Hyperammonaemia Page 6 of 22

7 Rhabdomyolysis / Metabolic Myopathies Condition Gene Original panel Primary transcript very long-chain acyl-coa dehydrogenase (VLCAD) deficiency ACADVL FAO NM_ Glycogen storage disease III AGL GSD NM_ Glycogen storage disease XII ALDOA GSD NM_ Muscular dystrophy, limb-girdle, type IC; Myopathy, distal, Tateyama type; Rippling muscle disease CAV3 Rhabdo NM_ CARNITINE PALMITOYLTRANSFERASE I, MUSCLE CPT1B FAO NM_ Carnitine palmitoyltransferase II (CPT II) deficiency CPT2 FAO NM_ Glycogen storage disease XIII ENO3 GSD NM_ MADD ETFAO FAO NM_ MADD ETFB FAO NM_ MADD (riboflavin-responsive) ETFDH FAO NM_ N/A - no mutations reported to date FBP2 GSD NM_ Glycogen storage disease II GAA GSD NM_ Glycogen storage disease IV / Polyglucosan body disease, adult form GBE1 GSD NM_ Glycogen storage disease XV GYG1 GSD NM_ Glycogen storage disease 0, muscle GYS1 GSD NM_ Mitochondrial TFP deficiency HADHA FAO NM_ Mitochondrial TFP deficiency HADHB FAO NM_ Hereditary myopathy with lactic acidosis ISCU Rhabdo NM_ Glycogen storage disease XI LDHA GSD NM_ Autosomal recessive recurrent acute myoglobinuria LPIN1 Rhabdo NM_ Glycogen storage disease VII PFKM GSD NM_ Glycogen storage disease X PGAM2 GSD NM_ Phosphoglycerate kinase 1 deficiency PGK1 GSD NM_ Glycogen storage disease XIV PGM1 GSD NM_ GSD Ixd; muscle phosphorylase kinase deficiency PHKA1 GSD NM_ PHKG1 GSD NM_ Glycogen storage disease, type V; McArdle disease PYGM GSD NM_ RBCK1 deficiency RBCK1 GSD NM_ Central Core Disease RYR1 Rhabdo NM_ Systemic primary carnitine deficiency (carnitine transporter) SLC22A5 FAO NM_ genes GSD - Glycogen Storage Diseases FAO - Fatty Acid oxidaton/metbaolism disorders Rhabdo - genes unique to rhabdomyolysis/metaboilc myopathies panel Page 7 of 22

8 Connective Tissue Disorders (SureSelect Design ID: ). Ehlers Danlos Syndrome (EDS) Subpanel Gene Primary transcript Vascular COL3A1 NM_ Classical COL5A1 NM_ COL5A2 NM_ Kyphoscoliotic B4GALNT7 NM_ CHST14 DSE FKBP14 PLOD2 PRDM5 RIN2 SLC39A13 ZNF469 TGFBR1 TGFBR2 NM_ NM_ NM_ NM_ NM_ NM_ NM_ NM_ NM_ NM_ Dermatosparaxis ADAMTS2 NM_ EDS-Arthrochalasic COL1A1 (Exons 6-10) NM_ COL1A2 (Exons 6-10) NM_ Familial Thoracic Aortic Aneurysm (FTAA) panel Condition Gene Primary transcript FTAA CBS NM_ EDS-Vascular COL3A1 NM_ Marfan s Syndrome FBN1 NM_ Beal s Syndrome FBN2 NM_ FTAA FLNA NM_ FTAA GATA5 NM_ FTAA MFAP5 NM_ FTAA MYH11 NM_ FTAA MYLK NM_ FTAA NOTCH1 NM_ FTAA PRKG1 NM_ FTAA SKI NM_ FTAA SLC2A10 NM_ FTAA SMAD3 NM_ FTAA SMAD4 NM_ FTAA TGFB2 NM_ Page 8 of 22

9 FTAA TGFB3 NM_ Loeys-Dietz Syndrome TGFBR1 NM_ Loey s Dietz Syndrome/Marfan s syndrome TGFBR2 NM_ FTAA SLC2A10 NM_ Osteogenesis Imperfecta (OI) panels Subpanels Gene Primary transcript Dominant, Extended COL1A1 NM_ Dominant, Extended COL1A2 NM_ Dominant IFITM5 (c.-14 only) NM_ Recessive, Extended BMP1 NM_ Recessive, Extended BMP1 NM_ Recessive, Extended CREB3L1 NM_ Recessive, Extended CRTAP NM_ Recessive, Extended FKBP10 NM_ Recessive, Extended IFITM5 full NM_ Recessive, Extended LEPRE1 (P3H1) NM_ Recessive, Extended OSX (SP7) NM_ Recessive, Extended P4HB NM_ Recessive, Extended PLOD2 NM_ Recessive, Extended PLS3 NM_ Recessive, Extended PPIB NM_ Recessive, Extended SEC24D NM_ Recessive, Extended SERPINF1 NM_ Recessive, Extended SERPINH1 NM_ Recessive, Extended SPARC NM_ Recessive, Extended TAPT1 NM_ Recessive, Extended TMEM38B NM_ Recessive, Extended WNT1 NM_ Recessive, Extended XYLT2 NM_ Familial Porencephaly (FP) panel Condition Gene Primary transcript Familial Porencephaly COL4A1 NM_ Familial Porencephaly COL4A2 NM_ Page 9 of 22

10 Alport Syndrome (AP) panel Condition Gene Primary transcript Alport Syndrome COL4A3 NM_ Alport Syndrome COL4A4 NM_ Alport Syndrome COL4A5 NM_ Bethlem Myopathy panel Condition Gene Primary transcript Bethlem Myopathy COL6A1 NM_ Bethlem Myopathy COL6A2 NM_ Bethlem Myopathy COL6A3 NM_ Cutis Laxa (CL) panel Condition Gene Primary transcript Cutis Laxa ATP6V0A2 NM_ Cutis Laxa ATP7A NM_ Cutis Laxa EFEMP2(FBLN4) NM_ Cutis Laxa ELN NM_ Cutis Laxa FBLN5 NM_ NM_ Cutis Laxa LTBP4 (NG_ ) Cutis Laxa PYCR1 NM_ Stickler Syndrome panel Condition Gene Primary transcript Stickler syndrome COL11A1 NM_ Stickler syndrome COL11A1 NM_ Stickler syndrome COL11A2 NM_ Stickler syndrome COL2A1 NM_ Stickler syndrome COL9A1 NM_ Stickler syndrome COL9A2 NM_ Stickler syndrome COL9A3 NM_ Page 10 of 22

11 TruSight Cancer Panels (Illumina Catalog No. TG ). Breast and Ovarian gene panel (Primary) Breast-ovarian cancer, familial, BRCA1 NM_ Breast-ovarian cancer, familial, BRCA2 NM_ Li-Fraumeni Syndrome (By request) TP53 NM_ Breast and Ovarian gene panel (Extended) Breast cancer susceptibility ATM NM_ Tumor predisposition syndrome BAP1 NM_ Breast-ovarian cancer, familial, BRCA1 NM_ Breast-ovarian cancer, familial, BRCA2 NM_ Breast cancer, early-onset BRIP1 NM_ Breast cancer, lobular CDH1 NM_ Breast cancer, susceptibility to CHEK2 NM_ Breast cancer, susceptibility to PALB2 NM_ Breast-ovarian cancer, familial, susceptibility to, RAD51C NM_ Breast-ovarian cancer, familial, susceptibility to, RAD51D NM_ Li-Fraumeni Syndrome TP53 NM_ Cowden Syndrome PTEN NM_ Peutz-Jeghers Syndrome STK11 NM_ Breast cancer PPM1D NM_ Nijmegen Breakage Syndrome NBN NM_ Colorectal Cancer gene panel (Primary - HNPCC) Colorectal cancer, hereditary nonpolyposis, type MLH1 NM_ Colorectal cancer, hereditary nonpolyposis, type MSH2 NM_ Colorectal cancer, hereditary nonpolyposis, type MSH6 NM_ Colorectal cancer, hereditary nonpolyposis, type 4 (By request) PMS2 NM_ Colorectal Cancer gene panel (Primary - FAP) Familial Adenamatous Polyposis APC NM_ Colorectal cancer, hereditary nonpolyposis, type MUTYH NM_ Page 11 of 22

12 Colorectal Cancer gene Panel (Extended) Familial Adenamatous Polyposis APC NM_ Polyposis, juvenile intestinal BMPR1A NM_ Colorectal cancer, hereditary nonpolyposis, type MLH1 NM_ Colorectal cancer, hereditary nonpolyposis, type MSH2 NM_ Colorectal cancer, hereditary nonpolyposis, type MSH6 NM_ Colorectal cancer, hereditary nonpolyposis, type MUTYH NM_ Colorectal cancer, hereditary nonpolyposis, type PMS2 NM_ Cowden Syndrome PTEN NM_ Polyposis, juvenile intestinal SMAD4 NM_ Peutz-Jeghers Syndrome STK11 NM_ Fanconi Anaemia Fanconi anaemia, complementation group A FANCA NM_ Fanconi anaemia, complementation group B FANCB NM_ Fanconi anaemia, complementation group C FANCC NM_ Fanconi anaemia, complementation group D BRCA2 NM_ Fanconi anaemia, complementation group D FANCD2 NM_ Fanconi anaemia, complementation group E FANCE NM_ Fanconi anaemia, complementation group F FANCF NM_ Fanconi anaemia, complementation group G FANCG NM_ Fanconi anaemia, complementation group I FANCI NM_ Fanconi anaemia, complementation group J BRIP1 NM_ Fanconi anaemia, complementation group L FANCL NM_ Fanconi anaemia, complementation group M FANCM NM_ Fanconi anaemia, complementation group N PALB2 NM_ Fanconi anaemia, complementation group O RAD51C NM_ Fanconi anaemia, complementation group P SLX4 NM_ Fanconi anaemia, complementation group Q ERCC4 NM_ Page 12 of 22

13 Neurogenetic Motor Disorders (SureSelect Design ID: ). Hereditary Spastic Paraparesis 88 gene panel Adrenoleukodystrophy, ALD ABCD1 NM_ Ataxia, spastic, 5, autosomal recessive, SPAX AFG3L2 NM_ Spastic paraplegia 9A, autosomal dominant, SPG9A ALDH18A1 NM_ Spastic paralysis, infantile onset ascending, IAHSP ALS2 NM_ Spastic paraplegia 63, SPG AMPD2 NM_ Pettigrew syndrome, PGS AP1S2 NM_ Spastic paraplegia 47, autosomal recessive, SPG AP4B1 NM_ Spastic paraplegia 51, autosomal recessive, SPG AP4E1 NM_ Spastic paraplegia 50, autosomal recessive, SPG AP4M1 NM_ Spastic paraplegia 52, autosomal recessive, SPG AP4S1 NM_ Spastic paraplegia 48, autosomal recessive, SPG AP5Z1 NM_ Argininemia ARG1 NM_ Spastic paraplegia 61, autosomal recessive, SPG ARL6IP1 NM_ Spastic paraplegia 66, SPG66 ARSI NM_ Spastic paraplegia 3A, autosomal dominant, SPG3A ATL1 NM_ Spastic paraplegia ATP2B4 NM_ Spastic paraplegia 26, autosomal recessive, SPG B4GALNT1 NM_ Spastic paraplegia 17, autosomal dominant, SPG BSCL2 NM_ Spastic paraplegia 55, autosomal recessive, SPG C12orf65 NM_ Spastic paraplegia 43, autosomal recessive, SPG C19orf12 NM_ Neuropathy, hereditary sensory, with spastic paraplegia, CCT5 NM_ Spastic paraplegia 73, autosomal dominant, SPG CPT1C NM_ Cerebrotendinous xanthomatosis, CTX CYP27A1 NM_ Spastic paraplegia 56, autosomal recessive, SPG CYP2U1 NM_ Spastic paraplegia 5A, autosomal recessive, SPG5A CYP7B1 NM_ Spastic paraplegia 28, autosomal recessive, SPG DDHD1 NM_ Spastic paraplegia 54, autosomal recessive, SPG DDHD2 NM_ Spastic paraplegia DNM2 NM_ Leukoencephalopathy with vanishing white matter, VWM EIF2B5 NM_ Spastic paraplegia 64, autosomal recessive, SPG ENTPD1 NM_ Spastic paraplegia 62, autosomal recessive, SPG62 ERLIN1 NM_ Spastic paraplegia 18, autosomal recessive, SPG ERLIN2 NM_ Spastic paraplegia 35, autosomal recessive, SPG FA2H NM_ Spastic paraplegia FARS2 NM_ Amyotrophic lateral sclerosis 11, ALS FIG4 NM_ Page 13 of 22

14 Hereditary Spastic Paraparesis 88 gene panel Spastic paraplegia 68, SPG68 FLRT1 NM_ Cerebral palsy, spastic quadriplegic, 1, CPSQ GAD1 NM_ Giant axonal neuropathy-1, GAN GAN NM_ Spastic paraplegia 46, autosomal recessive, SPG GBA2 NM_ Spastic paraplegia GCH1 NM_ Oculodentodigital dysplasia, ODDD GJA1 NM_ Spastic paraplegia 44, autosomal recessive, SPG GJC2 NM_ Tay-Sachs disease, TSD HEXA NM_ Spastic paraplegia 13, autosomal dominant, SPG HSPD1 NM_ Spastic paraplegia 74, autosomal recessive, SPG IBA57 NM_ Mental retardation, X-linked, syndromic, Claes-Jensen type, MRXSCJ KDM5C NM_ Spastic paraplegia 8, autosomal dominant, SPG KIAA0196 NM_ Spastic paraplegia 30, autosomal recessive, SPG KIF1A NM_ Spastic ataxia 2, autosomal recessive, SPAX KIF1C NM_ Spastic paraplegia 10, autosomal dominant, SPG KIF5A NM_ Spastic paraplegia KLC4 NM_ MASA syndrome, MASA L1CAM NM_ Spastic paraplegia LYST NM_ Spastic paraplegia MAG NM_ Spastic paraplegia 70, SPG70 MARS NM_ Spastic ataxia 3, autosomal recessive, SPAX MARS2 NM_ Spastic paraplegia MT-ATP6 NC_ Ataxia, spastic, 4, SPAX MTPAP NM_ Spastic paraplegia 6, autosomal dominant, SPG NIPA1 NM_ Spastic paraplegia 45, autosomal recessive, SPG NT5C2 NM_ Spastic paraplegia 67, SPG67 PGAP1 NM_ Spastic paraplegia 2, X-linked, SPG PLP1 NM_ Spastic paraplegia 39, autosomal recessive, SPG PNPLA6 NM_ Alzheimer disease, type 3, with spastic paraparesis and apraxia, AD PSEN1 NM_ Spastic paraplegia 69, SPG69 RAB3GAP2 NM_ Spastic paraplegia 31, autosomal dominant, SPG REEP1 NM_ Spastic paraplegia 72, autosomal recessive, SPG REEP2 NM_ Spastic paraplegia 12, autosomal dominant, SPG RTN2 NM_ Spastic ataxia, Charlevoix-Saguenay type, SACS SACS NM_ Amyotrophic lateral sclerosis 16, juvenile, ALS SIGMAR1 NM_ Allan-Herndon-Dudley syndrome, AHDS SLC16A2 NM_ Dystonia 9, DYT SLC2A1 NM_ Spastic paraplegia 42, autosomal dominant, SPG SLC33A1 NM_ Spastic paraplegia 4, autosomal dominant, SPG SPAST NM_ Page 14 of 22

15 Hereditary Spastic Paraparesis 88 gene panel Spastic paraplegia 11, autosomal recessive, SPG SPG11 NM_ Spastic paraplegia 20, autosomal recessive, SPG SPG20 NM_ Mast syndrome SPG21 NM_ Spastic paraplegia 7, autosomal recessive, SPG SPG7 NM_ Spastic paraplegia 49, autosomal recessive, SPG TECPR2 NM_ Spastic paraplegia 57, autosomal recessive, SPG TFG NM_ Spastic paraplegia 59, autosomal recessive, SPG59 USP8 NM_ Spastic ataxia 1, autosomal dominant, SPAX VAMP1 NM_ Spastic paraplegia 53, autosomal recessive, SPG VPS37A NM_ Neurodegeneration with brain iron accululation 5, NBIA WDR45 NM_ Spastic paraplegia 60, autosomal recessive, SPG60 WDR48 NM_ Spastic paraplegia 71, SPG71 ZFR NM_ Spastic paraplegia 15, autosomal recessive, SPG ZFYVE26 NM_ Spastic paraplegia 33, autosomal dominant, SPG ZFYVE27 NM_ Familial Amyotrophic Lateral Sclerosis with or without Frontotemporal Dementia 42 gene panel Primary lateral sclerosis, juvenile, PLSJ ALS2 NM_ Amyotrophic lateral sclerosis 9, ALS ANG NM_ Familial Amyotrophic Lateral Sclerosis ANXA11 NM_ Familial Amyotrophic Lateral Sclerosis ARHGEF28 NM_ Spinocerebellar ataxia 2, SCA ATXN2 NM_ Frontotemporal dementia and/or amyotrophic lateral sclerosis 2, FTDALS CHCHD10 NM_ Amyotrophic lateral sclerosis 17, ALS CHMP2B NM_ Cerebrotendinous xanthomatosis, CTX CYP27A1 NM_ Familial Amyotrophic Lateral Sclerosis DAO NM_ Neuropathy, distal hereditary motor, type VIIB, HMN7B DCTN1 NM_ Amyotrophic lateral sclerosis 19, ALS ERBB4 NM_ Familial Amyotrophic Lateral Sclerosis EWSR1 NM_ Amyotrophic lateral sclerosis 11, ALS FIG4 NM_ Amyotrophic lateral sclerosis 6, with or without frontotemporal dementia, ALS FUS NM_ Spastic paraplegia 46, autosomal recessive, SPG GBA2 NM_ Frontotemporal lobar degeneration with TDP43 inclusions, GRN related, GRN NM_ Amyotrophic lateral sclerosis 20, ALS HNRNPA1 NM_ Inclusion body myopathy with early-onset Paget disease with or without frontotemporal dementia 2, IBMPFD HNRNPA2B1 NM_ Frontotemporal Dementia, FTD MAPT NM_ Page 15 of 22

16 Familial Amyotrophic Lateral Sclerosis with or without Frontotemporal Dementia 42 gene panel Amyotrophic lateral sclerosis 21, ALS MATR3 NM_ Amyotrophic lateral sclerosis, susceptibility to, ALS NEFH NM_ Familial Amyotrophic Lateral Sclerosis NEK1 NM_ Amyotrophic lateral sclerosis 12, ALS OPTN NM_ Amyotrophic lateral sclerosis 18, ALS PFN1 NM_ Amyotrophic lateral sclerosis, susceptibility to, ALS PRPH NM_ Amyotrophic lateral sclerosis 4, juvenile, ALS SETX NM_ Amyotrophic lateral sclerosis 16, juvenile, ALS SIGMAR1 NM_ Amyotrophic lateral sclerosis 1, ALS SOD1 NM_ Spastic paraplegia 4, autosomal dominant, SPG SPAST NM_ Amyotrophic lateral sclerosis 5, juvenile, ALS SPG11 NM_ Spastic paraplegia 20, autosomal recessive, SPG SPG20 NM_ Frontotemporal dementia and/or amyotrophic lateral sclerosis 3, FTDALS SQSTM1 NM_ Familial Amyotrophic Lateral Sclerosis SS18L1 NM_ Familial Amyotrophic Lateral Sclerosis TAF15 NM_ Amyotrophic lateral sclerosis 10, with or without FTD, ALS10 Frontotemporal dementia and/or amyotrophic lateral sclerosis 4, FTDALS4 Amyotrophic lateral sclerosis 22 with or without frontotemoral dementia, ALS22 Amyotrophic lateral sclerosis 15, with or without frontotemporal dementia, ALS TARDBP NM_ TBK1 NM_ TUBA4A NM_ UBQLN2 NM_ Amyotrophic lateral sclerosis 8, ALS VAPB NM_ Amyotrophic lateral sclerosis 14, with or without frontotemporal dementia, ALS VCP NM_ Familial Amyotrophic Lateral Sclerosis VPS54 NM_ Pontocerebellar hypoplasia type 1A, PCH1A VRK1 NM_ Page 16 of 22

17 Dementia gene 27 gene panel Cerebral amyloid angiopathy, APP Related APP NM_ Alzheimer disease 1, familial, AD APP NM_ Frontotemporal dementia and/or amyotrophic lateral sclerosis 2, FTDALS CHCHD10 NM_ Frontotemporal Dementia, Chromosome 3-Linked, FTD CHMP2B NM_ Leukoencephalopathy, diffuse hereditary, with spheroids, HDLS CSF1R NM_ Cerebrotendinous xanthomatosis, CTX CYP27A1 NM_ Perry syndrome DCTN1 NM_ Neuropathy, hereditary sensory, type IE, HSN1E DNMT1 NM_ Amyotrophic lateral sclerosis 6, with or without frontotemporal dementia, ALS FUS NM_ Frontotemporal lobar degeneration with TDP43 inclusions, GRN related GRN NM_ Inclusion body myopathy with early-onset Paget disease with or without frontotemporal dementia 2, IBMPFD HNRNPA2B1 NM_ Cerebral arteriopathy, autosomal recessive, with subcortical infarcts and leukoencephalopathy, CARASIL HTRA1 NM_ Cerebral amyloid angiopathy, ITM2B-related, ITM2B NM_ Frontotemporal Dementia, FTD MAPT NM_ Amyotrophic lateral sclerosis 21, ALS MATR3 NM_ Cerebral arteriopathy with subcortical infarcts and leukoencephalopathy, CADASIL NOTCH3 NM_ Gerstmann-Straussler disease, GSD PRNP NM_ Alzheimer disease, type 3, AD PSEN1 NM_ Alzheimer disease-4, AD PSEN2 NM_ Mast syndrome SPG21 NM_ Frontotemporal dementia and/or amyotrophic lateral sclerosis 3, FTDALS SQSTM1 NM_ Amyotrophic lateral sclerosis 10, with or without FTD, ALS TARDBP NM_ Frontotemporal dementia and/or amyotrophic lateral sclerosis 4, FTDALS TBK1 NM_ Polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy, PLOSL TREM2 NM_ Amyotrophic lateral sclerosis 22 with or without frontotemoral dementia, ALS TUBA4A NM_ Polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy, PLOSL TYROBP NM_ Amyotrophic lateral sclerosis 15, with or without frontotemporal dementia, ALS UBQLN2 NM_ Amyotrophic lateral sclerosis 14, with or without frontotemporal dementia, ALS VCP NM_ Page 17 of 22

18 Spinal Muscular Atrophy 29 gene panel Charcot-Marie-Tooth disease, axonal, type 2N, CMT2N AARS NM_ Spinal muscular atrophy with progressive myoclonic epilepsy, SMAPME ASAH1 NM_ Spinal muscular atrophy, distal, X-linked 3, SMAX ATP7A NM_ Spinal muscular atrophy, lower extremity-predominant, 2, AD, SMALED BICD2 NM_ Spastic paraplegia 17, autosomal dominant, SPG BSCL2 NM_ Spinal muscular atrophy, Jokela type, SMAJ CHCHD10 NM_ Neuropathy, distal hereditary motor, type VIIB, HMN7B DCTN1 NM_ Spinal muscular atrophy, distal, autosomal recessive, 5, DSMA DNAJB2 NM_ Spinal muscular atrophy, lower extremity-predominant 1, AD, SMALED DYNC1H1 NM_ Pontocerebellar hypoplasia, type 1B, PCH1B EXOSC3 NM_ Pontocerebellar hypoplasia, type 1C, PCH1C EXOSC8 NM_ Neuronopathy, distal hereditary motor, type IID, HMN2D FBXO38 NM_ Neuropathy, distal hereditary motor, type VA, HMN5A GARS NM_ Tay-Sachs disease, TSD HEXA NM_ Neuropathy, distal hereditary motor, type IIB, HMN2B HSPB1 NM_ Neuronopathy, distal hereditary motor, type IIC, HMN2C HSPB3 NM_ Neuropathy, distal hereditary motor, type IIA, HMN2A HSPB8 NM_ Spinal muscular atrophy, distal, autosomal recessive, 1, DSMA IGHMBP2 NM_ Spinal muscular atrophy LAS1L NM_ Spinal muscular atrophy, distal, autosomal recessive, 4, DSMA PLEKHG5 NM_ Neuronopathy, distal hereditary motor, type VB, HMN5B REEP1 NM_ Cardioencephalomyopathy, fatal infantile, due to cytochrome c oxidase deficiency 1, CEMCOX SCO2 NM_ Spinal muscular atrophy, distal, autosomal recessive, 2, DSMA SIGMAR1 NM_ Neuronopathy, distal hereditary motor, type VIIA, HMN7A SLC5A7 NM_ Spinal muscular atrophy-1, SMA SMN1 NM_ Scapuloperoneal spinal muscular atrophy, SPSMA TRPV4 NM_ Spinal muscular atrophy, X-linked 2, infantile, SMAX UBA1 NM_ Spinal muscular atrophy, late-onset, Finkel type, SMAFK VAPB NM_ Pontocerebellar hypoplasia type 1A, PCH1A VRK1 NM_ Page 18 of 22

19 Neurogenetic Movement Disorders (SureSelect Design ID: ) Dystonia and Parkinsonism gene 27 gene panel Ataxia, spastic, 5, autosomal recessive; SPAX AFG3L2 NM_ Dystonia 24; DYT ANO3 NM_ Dystonia-12; DYT ATP1A3 NM_ Wilson disease; WD ATP7B NM_ Cerebrotendinous xanthomatosis; CTX CYP27A1 NM_ Spastic paraplegia 35, autosomal recessive; SPG FA2H NM_ Neurodegeneration with brain iron accumulation 3; NBIA FTL NM_ Parkinson disease, late-onset; PD GBA NM_ Dystonia, DOPA-responsive, with or without hyperphenylalaninemia; DRD GCH1 NM_ Dystonia 25; DYT GNAL NM_ Parkinson disease 8, autosomal dominant; PARK LRRK2 NM_ Dementia, frontotemporal, with or without parkinsonism; FTD MAPT NM_ Neurodegeneration with brain iron accumulation 1; NBIA1 (Pantothenate kinase-associated neurodegeneration; PKAN, PANK2 NM_ Parkinson disease, juvenile, type 2; PARK PARK2 NM_ Parkinson disease 7, autosomal recessive early-onset; PARK7 Parkinson disease 6, autosomal recessive early onset; PARK PARK7 NM_ PINK1 NM_ Paroxysmal nonkinesigenic dyskinesia 1; PNKD PNKD NM_ Spinocerebellar ataxia 14; SCA PRKCG NM_ Episodic kinesigenic dyskinesia 1; EKD1 (Dystonia 10; DYT10) PRRT2 NM_ Dystonia-11, myoclonic; DYT SGCE NM_ Allan-Herndon-Dudley syndrome; AHDS SLC16A2 NM_ Dystonia 9; DYT SLC2A1 NM_ Parkinson disease 1 autosomal dominant; PARK SNCA NM_ Spastic paraplegia 11, autosomal recessive; SPG SPG11 NM_ Dystonia, dopa-responsive, due to sepiapterin reductase deficiency; SPRD SPR NM_ Segawa syndrome, autosomal recessive TH NM_ Dystonia 6, torsion; DYT THAP1 NM_ Neurodegeneration with brain iron accumulation 5; NBIA WDR45 NM_ Page 19 of 22

20 Hereditary Ataxia and Migraine 42 gene panel Anaemia, sideroblastic and spinocerebellar ataxia; ASAT ABCB7 NM_ Ataxia, spastic, 5, autosomal recessive; SPAX5; Spinocerebellar ataxia 28; SCA AFG3L2 NM_ Ataxia, early-onset, with oculomotor apraxia and hypoalbuminemia; EAOH (AOA1) APTX NM_ Ataxia-telangiectasia; AT ATM NM_ Migraine, familial hemiplegic, 2; FHM ATP1A2 NM_ Cerebellar ataxia, areflexia, pes cavus, optic atrophy, and sensorineural hearing loss (CAPOS) ATP1A3 NM_ Wilson disease; WD ATP7B NM_ Spinocerebellar ataxia, infantile-onset; IOSCA C10orf2 NM_ Episodic ataxia, type 2; EA2; Migraine, familial hemiplegic, 1, with or without progressive cerebellar ataxia; FHM CACNA1A NM_ Episodic ataxia, type 5; EA CACNB4 NM_ Spastic paraplegia 56, autosomal recessive, SPG CYP2U1 NM_ Cerebrotendinous xanthomatosis; CTX CYP27A1 NM_ Spastic paraplegia 54, autosomal recessive, SPG DDHD2 NM_ Spinocerebellar ataxia 26; SCA EEF2 NM_ Spinocerebellar ataxia 27; SCA FGF14 NM_ Neurodegeneration with brain iron accumulation 3; NBIA FTL NM_ Friedreich ataxia (including Friedreich ataxia with retained reflexes); FRDA (FARR) FXN NM_ Parkinson disease, late-onset; PD GBA NM_ Spastic paraplegia 46, autosomal recessive; SPG GBA2 NM_ Spinocerebellar ataxia 18; SCA18 (Sensorimotor neuropathy with ataxia, autosomal dominant; SMNA) IFRD1 NM_ Spinocerebellar ataxia 15; SCA ITPR1 NM_ Episodic ataxia, type1 (including episodic ataxia/myokymia syndrome); EA KCNA1 NM_ Spinocerebellar ataxia 13; SCA KCNC3 NM_ Spinocerebellar ataxia 19; SCA KCND3 NM_ Spastic ataxia 4, autosomal recessive; SPAX MTPAP NM_ Spinocerebellar ataxia 23; SCA PDYN NM_ Spinocerebellar ataxia 14; SCA PRKCG NM_ Episodic kinesigenic dyskinesia 1; EKD1 (Dystonia 10; DYT10) PRRT2 NM_ Spastic ataxia, Charlevoix-Saguenay type; SACS SACS NM_ Migraine, familial hemiplegic, 3; FHM SCN1A NM_ Spinocerebellar ataxia, autosomal recessive 1; SCAR SETX NM_ Marinesco-Sjogren syndrome; MSS SIL1 NM_ Allan-Herndon-Dudley syndrome; AHDS SLC16A2 NM_ Episodic ataxia, type 6; EA SLC1A3 NM_ Page 20 of 22

21 Hereditary Ataxia and Migraine 42 gene panel Dystonia 9; DYT SLC2A1 NM_ Spastic paraplegia 7, autosomal recessive; SPG SPG7 NM_ Spinocerebellar ataxia 5; SCA SPTBN2 NM_ Spinocerebellar ataxia 35; SCA TGM6 NM_ Spinocerebellar ataxia 11; SCA TTBK2 NM_ Vitamin E, familial isolated deficiency of; VED TTPA NM_ Spastic ataxia 1, autosomal dominant; SPAX VAMP1 NM_ Spastic paraplegia 15, autosomal recessive, SPG ZFYVE26 NM_ Episodic Ataxia 6 gene panel Episodic ataxia, type 2; EA2; Migraine, familial hemiplegic, 1, with or without progressive cerebellar ataxia; FHM CACNA1A NM_ Episodic ataxia, type 5; EA CACNB4 NM_ Episodic ataxia, type1 (including episodic ataxia/myokymia syndrome); EA KCNA1 NM_ Episodic kinesigenic dyskinesia 1; EKD1 (Dystonia 10; DYT10) PRRT2 NM_ Episodic ataxia, type 6; EA SLC1A3 NM_ Dystonia 9; DYT SLC2A1 NM_ Hemiplegic Migraine 6 gene panel Migraine, familial hemiplegic, 2; FHM ATP1A2 NM_ Episodic ataxia, type 2; EA2; Migraine, familial hemiplegic, 1, with or without progressive cerebellar ataxia; FHM CACNA1A NM_ Episodic kinesigenic dyskinesia 1; EKD1 (Dystonia 10; DYT10) PRRT2 NM_ Migraine, familial hemiplegic, 3; FHM SCN1A NM_ Episodic ataxia, type 6; EA SLC1A3 NM_ Dystonia 9; DYT SLC2A1 NM_ Page 21 of 22

22 Bleeding & Thrombotic disorders (SureSelect Design ID: ) Sub-panels Disease Names Gene(s) OMIM Primary Transcript ADAMTS13 deficiency ADAMTS NM_ Factor V deficiency F NM_ Haemophilia A F NM_ Haemophilia A and B (where the type of haemophilia is unknown) F8 F NM_ NM_ Haemophilia A and 2N VWD (where the cause of reduced FVIII:C is unknown) F8 VWF NM_ NM_ Factor XIII deficiency F13A1 F13B NM_ NM_ Fibrinogen disorders FGA FGB FGG NM_ NM_ NM_ Glanzmann thrombasthenia ITGAT2B ITGB NM_ NM_ MYH9 related disorders MYH NM_ von Willebrand disease VWF NM_ Page 22 of 22