1. Carlos A. Saavedra-Matiz, MD NYSDOH-NBS Program DNA Lab NYMAC Albany, NY June 4-5, 2012 David Phillip Vetter (Sept 21, 1971 – Feb 22, 1984)Sept 211971Feb 221984

2. Immune System (IS) Function: Recognize and eliminate invading foreign organisms and molecules Peripheral Blood Lympochytes (IR or immune response): T Cells: Undergo differentiation in the thymus. Cellular IR B Cells: Undergo differentiation in the bone marrow. Humoral IR Lymphatic Organs: Primary: Thymus and bone marrow Secondary: Lymph nodes and spleen “An extremely efficient, genetically complex IS has evolved. Its development and function are regulated by genes of extraordinary diversity.” - Antibodies (Immunoglobulines) - Antigen-antibody binding - Antigen Receptors (TCR)

6. Multiple genotypes of SCID Gene defect Molecular pathogenesis % SCID cases * * T cells † † B cells Natural killer cells IL2RG (X-linked) Failure of IL-2 receptor (common) γ-chain signaling by cytokines IL-2, -4, - 7, -9, -15, -21 16 16 45-50 Low/ absent PresentAbsent ADAAdenosine deaminase deficiency 7 7 16 Low/ absent Absent IL7RFailure of signaling through IL-7 receptor 18 18 9 Low/ absent Present JAK3Failure of Janus kinase 3 activation by common γ-chain 17 17 6 Low/ absent PresentAbsent DCLRE1C ‡ ‡ Failure of antigen receptor rearrangement 19 19 <5Absent Present RAG1 ‡ ‡ Failure of antigen receptor rearrangement 19 19 <2Absent Present RAG2 ‡ ‡ Failure of antigen receptor rearrangement 19 19 <2Absent Present CD45 Lack of a cell surface protein tyrosine phosphatase receptor (PTPRC), required for T- and B-cell activation by antigen 20 and 21 2021 Rare Low/ absent Present Low/ variable TCRDT-cell receptor δ chain deficiency 22 22 Very rare Low/ absent Present LCK Lack of lymphocyte tyrosine kinase p56lck, required for T-cell development and activation 23 23 Very rare Low/ absent Present FOXN1 Lack of forkhead box N1transcription factor, required for thymus and hair follicle development (ortholog of nude mouse) 24 24 Very rare Low/ absent Present Deletion of Chr 22q11 Complete DiGeorge syndrome with thymus aplasia; part of DiGeorge- Velocardiofacial syndrome 25 25 RareAbsentVariablePresent Currently unknown Unknown, including reticular dysgenesis 26 and congenital anomaly syndromes with SCID 27 and 28 26 2728 Each one rare Low/ absent Variable

8. SAMPLES ARE PUNCHED INTO 96-WELL PLATES Wadsworth Center

10. Duplex Amplification Plots TREC Amplification plotRNase P amplification plot ADA

16. From Sept 29, 2010 to April 30, 2012: 382,406 infants screened No false negative cases

17. Overall PPV = 18.7% Classic SCID PPV = 2.1%

18. SCIDLeaky SCID/ Omenn Sy Variant SCID Syndromes with T cell impairment Secondary T cell lymphopenia Pre term alone IL2RG No Known Mut DiGeorge (22q11.2 deletion) Intestinal lymphangiectasia JAK3 CHARGEAnasarca IL7R JacobsenGastroschisis RAG1 RAC2 defectThird-spacing RAG2 DOCK 8 deficiency Gastrointestinal atresia ADA Ataxia telangiectasia Cardiac surgery +/- thymectomy CD45 VACTERL Association Congenital heart defects DCLRE1C Barth SyndromeNeonatal leukemia AK2OtherTAR SyndromeChylothorax OtherDown SyndromeChyloperitoneum Und (IL-7 Signaling Defect > 300 T cell impairmen t Ectrodactilia ectrodermal dysplasia Hypoplastic left heart defect Multiple congenital anomalies Degenerative neuromuscular Presumed metabolic dis.

19. The Ion Torrent Personal Genome Machine (PGM TM ) sequencer enables researchers to obtain highly accurate sequence in record time.

20. doi:10.1038/nature10242 RESEARCH SUPPLEMENTARY INFORMATION Figure S1 Process overview

21. GeneGene c.DNA PosRef PosAmplicon SizeRefIon SNPFreq 1Freq 2Counts 1Counts 2Ion CoverageSanger SNP% Concor danceComments MCAD-13754056702AA/G51.948.1329305634A/G100 MCAD-10284403702TT/C51.148.9505484989T/C100 MCAD-9854446702CG/C50.549.45165051021G/C100 MCAD-7254703779CG76.623.4441135576G/C50 MCAD-4095022779GG/A58.741.310171172G50Polymorphic region MCAD-3615069779TT0T/C50 MCAD-2575174779GA82.317.734574419A/G50 MCAD30+675527453CG72.427.6712798G/C50 MCAD708+7926637564AA/C62.038.08854142A50Primer Region MCAD708+8526643564TT/A60.639.48354137T50Primer Region MCAD116141980401AA/G53.646.1207178385A/G100p.V387V MCAD118442003401AA/G52.347.5232211443A/G100p.K395R GALC-1965252930TC/T58.641.49970169C/T100 GALC-75441930GG/C52.048.09083173G/C100 GALC135460312GG/C52.247.06054114G/C100p.A5P GALC275474312CA/C62.236.7613697A/C100p.G9G GALC148-709971357GA/G55.944.16652118G50Polymorphic region GALC148-439998357CT100.0178 C0Polymorphic region GALC281-3511927344GG/A55.444.6128103231G/A100 GALC28611967344AA/G50.0 126 252A/G100p.T96A GALC30111982344AG/A52.547.5124112236G/A100p.M101V GALC395-5113980421GA/G51.248.8222143A/G100 GALC534+8914269421CC/T60.539.5463076C/T100 GALC69422196526GA/G63.636.47411A/G100p.D232N GALC704+5622262526TT/C55.644.4549T/C100 GALC705-5930015564CT/C50.849.2130126256T/C100 GALC1290+2348742343TC68.631.416877245C/T50 GALC1291-6150625404CC/T53.446.6127111238C/T100 GALC157252961417GG/A51.748.3138129267G/A100P.T524T (Cis-M101V) GALC1623-1556991416CT/C51.748.39084174T/C100 GALC165057033416AA/T50.549.59997196A/T100p.V550V GALC1786+557174416CC/G58.042.09468162C/G100Cis-M101V GALC187363695570AG/A54.944.8197161358G/A100p.A625T (Cis-M101V) VLCAD351541560GA/G55.144.97561136A/G100p.M1I VLCAD138+9954631560TG/T54.145.96051111G/T100 VLCAD477+5262771560AT67.932.1552681A50 VLCAD478-10166041061AC77.522.57923102C100? VLCAD623-871111061CT65.834.212766193T/C50 VLCAD160095551040GG/A51.848.2177165342G/A100p.E534K VLCAD1605+595661040TT/C51.948.1180167347T/C100 PKU168+199831291TC/T56.443.37745941368C/T100 PKU353-2245031340CC/T50.449.6185182367C/T100 PKU442-4855892287AG100.0211 G100 PKU510-5467217354GA/G52.247.85655181083A/G100 PKU69667457354AA/G61.438.67844931277A/G100p.Q312Q PKU73569681423GG/A54.046.05935051098G/A100p.V245V PKU115578913264GC/G50.949.1122911842413C/G100p.L383L GALT1-285040413TA99.1 316 A100 GALT8557395570GG/T53.546.1251216467G/T100p.K285N GALT9597827459CC/T55.344.7198160358C/T100p.A320V GALT*1248936360AT100.0 693 T100