NR0B1 Supporting Information Biorxiv

Home , CREB3, HSD17B3, IL1RAPL1, PHOX2B, SF1 (gene), SIX3

Supporting Information:

Supporting Figures

Figure S1. Comparison of Xp21.2 CNVs. Schematic representation comparing NR0B1 locus copy number variations in patients presented in this study with those previously described in the literature and associated with 46, XY sex reversal. The green bars represent the duplications and the triplication described in the patients of this study (P1 -P3) and the blue bars represent duplications previously reported by others researchers (Barbaro, Oscarson et al. 2007, Barbaro, Cicognani et al. 2008, Ledig, Hiort et al. 2010, White, Ohnesorg et al. 2011, Barbaro, Cook et al. 2012, Dong, Yi et al. 2016, Garcia-Acero, Molina et al. 2019) . All previously reported duplications include the NR0B1 (red underlined), CXorf21 and GK gene. The red bar marks the deletion reported by Smyk et al. (Smyk, Berg et al. 2007). There is a mutual 35kb region present in all Xp21.2 copy number variations associated with 46,XY GD, highlighted by the green box. The representation was generated using the UCSC Genome Browser (GRCh37/hg19)(Kent, Sugnet et al. 2002).

Figure S2. Sox9 activation during testicular development. Mechanism of Sox9 regulation, as suggested by Ludbrook et al. (2012). Sox9 gene with upstream enhancer Enh13 as grey bar. Grey spheres symbolize SF1 and SRY/SOX9 binding sites within the enhancer. Besides Enh13 (Gonen, Futtner et al. 2018) also an enhancer termed TESCO (Sekido and Lovell-Badge 2008) has previously been proposed as especially relevant for testis specific Sox9 expression in mice. Sox9 enhancers appear to be activated in three phases (i-iii) (Sekido and Lovell-Badge 2008). The individual enhancers may have varying phases of predominant activation and relevance for sex determination. (i) Sox9 expression is initiated by SF1, independently of SRY. (ii) Following initiation, Sox9 is upregulated synergistically by SRY and SF1. Sry is only transiently expressed in mice (Sekido, Bar et al. 2004), due to repression by SOX9 (Chaboissier, Kobayashi et al. 2004). (iii) Sox9 enhancer activation is maintained in SRY absence through autoregulation by SOX9. SOX9 substitutes SRY as they share the same DNA-Binding motif and it also synergizes with SF1. Thus, SRY is only briefly needed to increase Sox9 expression above a threshold allowing self-sustained expression (Sekido and Lovell-Badge 2009). If this threshold is not reached in time, this leads to the commitment of the supporting cell line precursors to the ovary determining pathway through accumulating ovary-determining/anti- testis factors (Gonen, Futtner et al. 2018). It is of note however, that in humans SOX9 enhancer duplications were found in several 46,XX SRY-negative males. This manifestation is attributed to enhancers termed eSR-A (enhancer sex reversal A) and eSR-B, which seem to be predominantely activated through SOX9 and SF1 (Croft, Ohnesorg et al. 2018). SOX9 is essential for Sertoli cell differentiation and activates downstream targets including Fgf9/Fgfr2 and Sox8 (Sekido and Lovell- Badge 2008, Ludbrook, Bernard et al. 2012). NR0B1 (DAX1) interferes with SF1 activation of target genes possibly through protein-protein interactions (Nachtigal, Hirokawa et al. 1998) or displacement from DNA binding sites in a dose-dependent manner (Ludbrook, Bernard et al. 2012) SRY is insufficient to activate Sox9 expression alone, thus the necessary threshold to induce Sox9 autoregulation and subsequent testis formation is not reached (Sekido and Lovell-Badge 2008).

Figure S3: Sex development genes in patients 1 & 2 show intermediate expression between ovary and testis tissue. The combined beeswarm and box-whisker plots depict the log2-transformed transcript per million (TPM) expression for selected sex development genes of the GTEx ovary (left) and testis (middle) data (Consortium 2013, Carithers, Ardlie et al. 2015). Expression in the two patients is depicted on the right. The boxes and thick black bar denote the first to third quartile of the data and the whiskers extend the box by 1.5 of the interquartile range. A moderated t-test shows significant differential regulation of genes between all groups.

Figure S4. MLPA and CMA Analysis of P1. (A) MLPA of P1 revealing the duplication of CXorf21 probes at Xp21.2 [30,595,621-30,615,321] (GRCh37/hg19); signals for NR0B1, SRY, SOX9 and NR5A1 are normal. (B) Normal probe signals for DMRT1, CYP17A1, SRD5A2 and HSD17B3 genes in MLPA. (C) Chromosome micro array (CMA) analysis using software Chromosome Analysis Suite (ChAS - Affymetrix®) revealing a ≈277kb duplication of Xp21.2 (arr[GRCh37] Xp21.2(30580693_30857187)x2) covered by 165 markers. The minimal duplicated region includes the fully duplicated GK gene and parts of TAB3 and CXorf21.

Figure S5. aCGH and GS comparison in P2 at NR0B1 locus (A) aCGH results for P2. Green dots indicate probes used for hybridization. Results show two separate duplications as illustrated by increased hybridization of probes and emphasized by green bars. To the left a 389kb duplication (chrX:29,924,420-30,313,761; GRCh37/hg19) was detected by increased hybridization of 17 probes. The second duplication was detected as 447kb of size (chrX: 30,401,819-30,848,988) indicated by 23 probes. The duplications are separated by four probes labelled 1 – 4. Probes 1, 3 and 4 hybridized only to a single copy. Probe 2 showed no hybridization. (B) Probes 1 to 4 have been compared to genome sequencing data. For visualisation of WGS data of P2 (region ChrX:30,292,562-30,423,695) the Integrative Genome Viewer (IGV) was used (Robinson, Thorvaldsdottir et al. 2011). Increased number of reads (grey bars) at both sides of the viewing window signify duplication detected by WGS (green bars). Deletions are evident through the lack of reads in the two regions marked with red bars. Single hybridized CGH probes 1, 3 and 4 map to areas not duplicated according to WGS data. Probe 2 maps to a deletion and correspondingly did not hybridize.

Figure S6. aCGH in P3 at NR0B1 locus. Green dots indicate probes used for hybridization. Results show a 1,2 Mb triplication (ChrX:29,851,537-31,069,736) emphasized by green bar. The genes MAGEB1-4, NR0B1, CXorf21 and TAB3 are fully triplicated whereas IL1RAPL1 is only partially triplicated.

Figure S7. Structural Variation of Patient 3 including proposed TAD structure at NR0B1 locus. Genomic distances are not to scale. The chromosome segment is drawn with the distal chromosome arm to the left and centromere to the right. WGS showed the triplication originally mapped to chrX: 29,849,782 – 31,088,713 and is arranged in a tandem manner. * marks two identical 49 bp inserts at two of the breakpoints. The sequence originates from an intron of the DMD gene, located to the centromere of the SV event. Grey arrow heads represent binding sites of the insulator CCCTC-binding factor (CCTF). The arrangement is such, that between every NR0B1 gene and closest putative enhancer there is an insulator (CTCF) binding site. However, these sites only perform insulation when bound to a partnering CTCF. In lack of sufficient number of partnering CTCFs we propose three different possible TAD conformations (dashed grey lines). Each conformation enables NR0B1 enhancer adoption in different sets of gene/enhancer pairs (arrows).

Figure S8. NR0B1 TAD conformation of different human cell lines according to PennState 3D GenomeBrowser data. Green bars represent duplications of patients described in this study (P1 & P2). The red bar shows the deletion reported by Smyk et al. (2007). The TAD encompassing NR0B1 (NR0B1 TAD) and

8 the adjacent centromeric and telomeric TADs identified by Hi-C sequence data and made available through the PennState 3D Genome Browser (Wang et al., 2018) have been extracted for 36 cell lines. TAD position and size is visualized as blue bars for corresponding cell lines using the UCSC Genome Browser (GRCh37/hg19) (Kent, Sugnet et al. 2002). Most cell lines show a conserved NR0B1 TAD boundary at CXorf21, which is in a region shared by the duplications of P1 and P2 as well as the deletion reported by Smyk et al. (2007).

Figure S9. Binding sites in mutual region between P1 - P2 duplications, P3 triplication and the deletion reported by Smyk et al. (2007). Minimal region of overlap (35kb) between duplication of P1 reported in this study (green bars) and the deletion reported by Smyk et al. (2007) (red bar). This consensus region also covered by P2 and P3 encompasses the whole CXorf21 gene. ENCODE Chip-seq Data (Consortium 2012, Davis, Hitz et al. 2018) available in the UCSC genome browser (Kent, Sugnet et al. 2002) shows a CTCF binding site in this region together with binding sites for the cohesin complex components RAD21 and SMC3. We propose this to be the centromeric NR0B1 TAD border.

10 A B S PERMATOGENES IS 0.6 PHOX2B WNT BETA CATENIN SIGNALING GMNN LMX1A EPITHE LIAL MES ENCHYMAL TRANSITION 0.4 NOTCH S IGNALING SIX3 0.5 E E

TGF BETA S IGNALING MEIS2 n n r i c 0.2 r

UV RESPONSE DN i TCF15 c h h m

ADIPOGENESIS m E2F6 e e n n P53 PATHWAY 0 ZNF143 0 t S t S

MYOGE NESIS NKX6.1 c c o o r

RFX2 e

PI3K AKT MTOR S IGNALING r -0.2 e IL2 STAT5 S IGNALING CREB3 TNFA SIGNALING VIA NFKB PHOX2A -0.5 INTERFERON ALPHA RES PONSE -0.4 FOSL2.JUN IL6 J AK STAT3 S IGNALING HOXB9 INTERFERON GAMMA RES PONSE TBX3 -0.6 ANGIOGENES IS DLX2 COAGULATION MAF APICAL J UNCTION TEAD4 HYPOXIA NR1I3 APOPTOSIS AHR PROTEIN S ECRETION EPAS1 OXIDATIVE PHOSPHORYLATION PRDM1 MYC TARGETS V1 SMAD1 UNFOLDED PROTE IN RESPONSE TBX2 MYC TARGETS V2 DBP UV RESPONSE UP DLX5 G G G G G G G P P G T T T T T T e e e e e e a a

T T T T T T T HLF T s s s s s s t t E E E E E E E i i t t t t t t E e e i i i i i i s s s s s s x x x x x x x n n

x FOSL2 ------1 1 1 1 1 1 y y y S y o t t M 2 1 1 1 1 1 1 1 oung oung oung oung ld M e e E G D I E P e 78 ni KLF11 di e X r M 81 M S M o a - d 1 X M A A A r C 3 P - n 9 id 1 i - - a dul dul dul 1

1 HOXA5 2 id 52 - - 1 A O 0 72 4 6 n 7 A 2 g 26 - 6 v t t t S 2 26 e 6 g T a -

S NCOA2 6 - e - M S S es r - - S S y M - M M 5 M M - 9

5 TFEB t - - i 5 5 87 P - - s 5 5 N G H A 9 8

9 TEAD1 I G 1 5 D D 1 K S U 1 P V TWIST1 GTEx-11EM3-1726-SM-5N9D1 GTEx-11I78-1926-SM-59878 GTEx-11P81-1526-SM-5P9GS GTEx-11GSP-0226-SM-5A5KV GTEx-11DXX-1426-SM-5GIDU Ovary Median GTEx GTEx-11EMC-1726-SM-5H11P olderMidAge Testis Senior Testis youngAdult Testis youngAdult Testis youngAdult Testis Testis Median GTEx youngMidAge Testis 2 Patient 1 Patient

1 adj. P-value [-log10]

CELL CYCLE

CELL CYCLE MITOTIC CELL CYCLE CHECKPOINTS

Figure S10. Patient transcriptomes display intermediate pathway activity between testis and ovary tissue. The heatmap depicts a gene set variation analysis (GSVA) on all hallmark gene sets (A) and upstream transcription factors (B) that differ significantly according to a moderated F-statistic between the three groups ovary, testis and patient samples (adjusted p-value < 0.01 and < 0.001 in A and B). The sample groups consist of seven random GTEx ovary tissues and adult testis tissues from the ArrayExpress E-MTAB-6814 data (Athar, Fullgrabe et al. 2019) as well as the GTEx median tissue expression and the two patient samples. Colors encode the per sample enrichment score of the GSVA. Columns and rows have been hierarchically clustered by a complete linkage method according to their Euclidean distance. (C) P-value significance of downregulation of Reactome cell cycle pathway in the patient samples relative to the testis and ovary samples. Gene sets have been extracted from the C2 curated gene sets from MSigDB (Liberzon, Birger et al. 2015).

Figure S11. Overview of structural variations in P1 and P2. Diagram compares the arrangements in patient 1 (B) and patient 2 (C) with the reference (A) genomic arrangement. Chromosomal positions are given according to GRCh37/hg19.

Figure S12. GeneHancer Promoters and Enhancers upstream of NR0B1. Data from the GeneHancer database (Fishilevich, Nudel et al. 2017) was visualized in the UCSC genome browser (Kent, Sugnet et al. 2002). Promoters and enhancers are distinguished by red and grey colours respectively in the GH Reg Elements (DE) track. Colour intensity shows confidence score of the respective enhancers. According to Fishilevich et al. (2017) this is based on: “the number of supporting data sources, the number of unique TFBSs contained, and the overlap with ultra-conserved non-coding genomic elements“. Arcs in this tack show proposed gene-enhancer interaction in the reference genome.

14 Figure S13. Proposed structures of topologically associating domain (TAD) at NR0B1 locus. (A) Reference sequence for comparison with patients with structural variations (B)(C)(D). Arrow heads represent binding sites of CCCTC-binding factor (CCTF) which can function as an insulator and TAD border. When convergent oriented, CTCFs can form a TAD as represented by the grey triangle. (B) We suggest the possibility of two different TAD conformations in P1 as indicated by the continuous and dashed grey triangle. Only one of the conformations can be present at a time. The conformation depicted by the continuous grey triangle would more readily allow interaction between enhancers (grey spheres) located between GK and CXorf21 with the NR0B1 promoter, as these would then be located in a mutual TAD. (C) Correspondingly the inversion in P2 brings these non-cognate enhancers into the close proximity to NR0B1 while evading the shielding effect of a TAD border between the gene and non-cognate enhancers. (D) The deletion reported by Smyk et al. (2007) disrupts the conventional TAD conformation through CTCF motif deletion, thus allowing increased interactions between non-cognate enhancers and the NR0B1 promoter

15 Supporting Tables

Table S1. Mutations in Genes Associated With 46,XY DSD Found in Patients 1 and 2.

Patient Gene dbSNP ID ExAc All ExAc SIFT Polyphen2 MutationTaster cDNA change Protein Change (Lek et European (Vaser et (Adzhubei et (Schwarz et al., al., 2016) Non-Finish al., 2016) al., 2010) 2014) (Lek et al., 2016) P1 STAR rs34908868 0.00158 0.00249 D (0.009) D (1.0) D (0.996) NM_000349.2:c.361C>T NP_000340.2:p.Arg121Trp

P1 POR rs145782750 0.00174 0.00451 T (0.904) B (0.156) D (1.0) NM_000941.2:c.1891G>A NP_000932.3:p.Val631Ile

P2 ZFPM2 rs202217256 0.00366 0.00567 T (0.422) P (0.914) D (1.0) NM_012082.3:c.292G>A NP_036214.2:p.Asp98Asn

P3 ESR2 rs367855747 0.0002 0.0004 D (0.021) D (0.975) D (0.999) NM_001214902.1:c. NP_001201831.1:

1331G>A p.Ser444Asn

All three mutations are heterozygous missense SNVs.

SIFT: D = Deleterious, T = Tolerated; Polyphen2: B = benign, D = probably damaging, P = possibly damaging; MutationTaster: D = disease causing

16 Table S2. Primers for PCR and Sanger Sequencing in Patients 1 and 2

Patient Breakpoint Direction Primer sequence 5’to 3’ Annealing (BP) Temperature (°C) P1 BP 1 Fwd CAA ATG CTG GTG GGG TTA TG 55.5 Rev TCA AAT TGG CTT CCT TCT CTT AGT 55.5 BP 2 Fwd TTT CCA ATG CAC TCC CTT AGA CAG 55.5 Rev CCT TTT CAA ATT GGC TTC CTT C 55.5 BP 3 Fwd GTT TGT CTG GCG ATA CTT CTT 55.5 Rev GTT TCA TCT GAT CTA CAT TTA 55.5 P2 PB 1 Fwd CCT TTT CAA ATT GGC TTC CTT C 58.1 Rev GGC ATC TTA CCA CCT CTG A 58.1 BP 2 Fwd TTC CCA ATT TCA ATC CTC ATA 50.3 Rev GAA ATC CAC TCT GCC AAC AC 50.3 BP 3 Fwd TAG TAC ATG GCA TTA TTT ATT GAC 58.1 Rev ATG GCA GTT GTA GGT AGA GAA GTA 58.1 BP 4 Fwd CAC CTT GCA TAT GGA AAA ACC 50.3 Rev TGA CTA GAC CAA TAA CAG GAT 50.3 In P1 also the transition from the duplicated to non-duplicated region downstream of CXorf21 and within TAB3 (above denoted as breakpoint 1 and 3) were sequenced. These sequences are not explicitly shown in Figure 1.

17 References:

Athar, A., A. Fullgrabe, N. George, H. Iqbal, L. Huerta, A. Ali, C. Snow, N. A. Fonseca, R. Petryszak, I. Papatheodorou, U. Sarkans and A. Brazma (2019). "ArrayExpress update - from bulk to single-cell expression data." Nucleic Acids Res 47(D1): D711-D715. Barbaro, M., A. Cicognani, A. Balsamo, A. Lofgren, L. Baldazzi, A. Wedell and M. Oscarson (2008). "Gene dosage imbalances in patients with 46,XY gonadal DSD detected by an in-house-designed synthetic probe set for multiplex ligation-dependent probe amplification analysis." Clin Genet 73(5): 453-464. Barbaro, M., J. Cook, K. Lagerstedt-Robinson and A. Wedell (2012). "Multigeneration Inheritance through Fertile XX Carriers of an NR0B1 (DAX1) Locus Duplication in a Kindred of Females with Isolated XY Gonadal Dysgenesis." Int J Endocrinol 2012: 504904. Barbaro, M., M. Oscarson, J. Schoumans, J. Staaf, S. A. Ivarsson and A. Wedell (2007). "Isolated 46,XY gonadal dysgenesis in two sisters caused by a Xp21.2 interstitial duplication containing the DAX1 gene." J Clin Endocrinol Metab 92(8): 3305-3313. Carithers, L. J., K. Ardlie, M. Barcus, P. A. Branton, A. Britton, S. A. Buia, C. C. Compton, D. S. DeLuca, J. Peter-Demchok, E. T. Gelfand, P. Guan, G. E. Korzeniewski, N. C. Lockhart, C. A. Rabiner, A. K. Rao, K. L. Robinson, N. V. Roche, S. J. Sawyer, A. V. Segre, C. E. Shive, A. M. Smith, L. H. Sobin, A. H. Undale, K. M. Valentino, J. Vaught, T. R. Young, H. M. Moore and G. T. Consortium (2015). "A Novel Approach to High-Quality Postmortem Tissue Procurement: The GTEx Project." Biopreserv Biobank 13(5): 311- 319. Chaboissier, M. C., A. Kobayashi, V. I. Vidal, S. Lutzkendorf, H. J. van de Kant, M. Wegner, D. G. de Rooij, R. R. Behringer and A. Schedl (2004). "Functional analysis of Sox8 and Sox9 during sex determination in the mouse." Development 131(9): 1891-1901. Consortium, E. P. (2012). "An integrated encyclopedia of DNA elements in the human genome." Nature 489(7414): 57-74. Consortium, G. T. (2013). "The Genotype-Tissue Expression (GTEx) project." Nat Genet 45(6): 580- 585. Croft, B., T. Ohnesorg, J. Hewitt, J. Bowles, A. Quinn, J. Tan, V. Corbin, E. Pelosi, J. van den Bergen, R. Sreenivasan, I. Knarston, G. Robevska, D. C. Vu, J. Hutson, V. Harley, K. Ayers, P. Koopman and A. Sinclair (2018). "Human sex reversal is caused by duplication or deletion of core enhancers upstream of SOX9." Nat Commun 9(1): 5319. Davis, C. A., B. C. Hitz, C. A. Sloan, E. T. Chan, J. M. Davidson, I. Gabdank, J. A. Hilton, K. Jain, U. K. Baymuradov, A. K. Narayanan, K. C. Onate, K. Graham, S. R. Miyasato, T. R. Dreszer, J. S. Strattan, O. Jolanki, F. Y. Tanaka and J. M. Cherry (2018). "The Encyclopedia of DNA elements (ENCODE): data portal update." Nucleic Acids Res 46(D1): D794-D801.

18 Dong, Y., Y. Yi, H. Yao, Z. Yang, H. Hu, J. Liu, C. Gao, M. Zhang, L. Zhou, Asan, X. Yi and Z. Liang (2016). "Targeted next-generation sequencing identification of mutations in patients with disorders of sex development." BMC Med Genet 17: 23. Fishilevich, S., R. Nudel, N. Rappaport, R. Hadar, I. Plaschkes, T. Iny Stein, N. Rosen, A. Kohn, M. Twik, M. Safran, D. Lancet and D. Cohen (2017). "GeneHancer: genome-wide integration of enhancers and target genes in GeneCards." Database (Oxford) 2017. Garcia-Acero, M., M. Molina, O. Moreno, A. Ramirez, C. Forero, C. Cespedes, J. C. Prieto, J. Perez, F. Suarez-Obando and A. Rojas (2019). "Gene dosage of DAX-1, determining in sexual differentiation: duplication of DAX-1 in two sisters with gonadal dysgenesis." Mol Biol Rep. Gonen, N., C. R. Futtner, S. Wood, S. A. Garcia-Moreno, I. M. Salamone, S. C. Samson, R. Sekido, F. Poulat, D. M. Maatouk and R. Lovell-Badge (2018). "Sex reversal following deletion of a single distal enhancer of Sox9." Science 360(6396): 1469-1473. Kent, W. J., C. W. Sugnet, T. S. Furey, K. M. Roskin, T. H. Pringle, A. M. Zahler and D. Haussler (2002). "The human genome browser at UCSC." Genome Res 12(6): 996-1006. Ledig, S., O. Hiort, G. Scherer, M. Hoffmann, G. Wolff, S. Morlot, A. Kuechler and P. Wieacker (2010). "Array-CGH analysis in patients with syndromic and non-syndromic XY gonadal dysgenesis: evaluation of array CGH as diagnostic tool and search for new candidate loci." Hum Reprod 25(10): 2637-2646. Liberzon, A., C. Birger, H. Thorvaldsdottir, M. Ghandi, J. P. Mesirov and P. Tamayo (2015). "The Molecular Signatures Database (MSigDB) hallmark gene set collection." Cell Syst 1(6): 417-425. Ludbrook, L. M., P. Bernard, S. Bagheri-Fam, J. Ryan, R. Sekido, D. Wilhelm, R. Lovell-Badge and V. R. Harley (2012). "Excess DAX1 leads to XY ovotesticular disorder of sex development (DSD) in mice by inhibiting steroidogenic factor-1 (SF1) activation of the testis enhancer of SRY-box-9 (Sox9)." Endocrinology 153(4): 1948-1958. Nachtigal, M. W., Y. Hirokawa, D. L. Enyeart-VanHouten, J. N. Flanagan, G. D. Hammer and H. A. Ingraham (1998). "Wilms' tumor 1 and Dax-1 modulate the orphan nuclear receptor SF-1 in sex- specific gene expression." Cell 93(3): 445-454. Robinson, J. T., H. Thorvaldsdottir, W. Winckler, M. Guttman, E. S. Lander, G. Getz and J. P. Mesirov (2011). "Integrative genomics viewer." Nat Biotechnol 29(1): 24-26. Sekido, R., I. Bar, V. Narvaez, G. Penny and R. Lovell-Badge (2004). "SOX9 is up-regulated by the transient expression of SRY specifically in Sertoli cell precursors." Dev Biol 274(2): 271-279. Sekido, R. and R. Lovell-Badge (2008). "Sex determination involves synergistic action of SRY and SF1 on a specific Sox9 enhancer." Nature 453(7197): 930-934. Sekido, R. and R. Lovell-Badge (2009). "Sex determination and SRY: down to a wink and a nudge?" Trends Genet 25(1): 19-29.

19 Smyk, M., J. S. Berg, A. Pursley, F. K. Curtis, B. A. Fernandez, G. A. Bien-Willner, J. R. Lupski, S. W. Cheung and P. Stankiewicz (2007). "Male-to-female sex reversal associated with an approximately 250 kb deletion upstream of NR0B1 (DAX1)." Hum Genet 122(1): 63-70. White, S., T. Ohnesorg, A. Notini, K. Roeszler, J. Hewitt, H. Daggag, C. Smith, E. Turbitt, S. Gustin, J. van den Bergen, D. Miles, P. Western, V. Arboleda, V. Schumacher, L. Gordon, K. Bell, H. Bengtsson, T. Speed, J. Hutson, G. Warne, V. Harley, P. Koopman, E. Vilain and A. Sinclair (2011). "Copy number variation in patients with disorders of sex development due to 46,XY gonadal dysgenesis." PLoS One 6(3): e17793.