Figure S2 The pedigree chart of all sampled Fleckvieh bulls composing case and carrier group in Table S1. The case individuals (PP) are represented by solid circles (females) and squares (males); declared carriers by half-filled symbols; not sampled individuals are marked with a diagonal line. To reduce complexity of the pedigree not all relationships were presented. At two positions (A and B) there are relationships to important sires indicated. The inbreeding of the Fleckvieh bull (PP-DFV1) chosen for re-sequencing is superimposed by red lines. All sampled Fleckvieh PP animals bear two copies of the common haplotype (AGACAAGGA) and were genotyped as PC/PC. All Pp bear one copy of the common haplotype and were genotyped as PC/prs. No recombination was detected in the common haplotype. (TIF) Figure S3 The pedigree chart of all sampled Holstein bulls composing case and carrier group in Table S1. The case individuals (PP) are represented by solid circles (females) and squares (males); declared carriers by half-filled symbols; not sampled individuals are marked with a diagonal line. To reduce complexity of the pedigree not all relationships were presented. At positions A, B and C there are relationships to important sires indicated. The founder of the Celtic polledness is indicated by D. The carriers and cases of Friesian and Celtic polledness are filled with gray (genotyped as PF/prs and PF/PF) and black color (genotyped as PC/prs and PC/PC), respectively. Two heteroge- neous polled bulls were genotyped as PC/PF. At the SNP-Chip level all sampled Holstein PP and Pp animals bear two or one copy of the common haplotype (AGACAAGGA) and there was no recombination detected. (TIF) Figure S4 The pedigree chart of all sampled case and carrier Jersey bulls. The case individuals (PP) are represented by solid circles (females) and squares (males); declared carriers (Pp) by half-filled symbols; not sampled individuals are marked with a diagonal line. The founder of the Friesian polledness in Jersey breed is indicated by F. The carriers and cases of polledness with Friesian and Celtic origin are filled with gray (genotyped as PF/prs and PF/PF) and black color (genotyped as PC/prs and PC/PC), respectively. Two heterogeneous polled bulls were genotyped as PC/PF. At the SNP-Chip level all sampled Jersey PP and Pp animals bear two or one copy of the common haplotype (AGACAAGGA) and there was no recombination detected. (TIF) Figure S5 Across-species conservation for variant PC1655463T. (a) The sequence conservation around the position of the candidate mutation PC1655463T among animals without horn, horn-bearing and among all animals are represented as PhastCons scores, calculated from the underlying multi-species alignment in plot b. (b) Sequence identity among horn-bearing animals is denoted with red stars and black stars are used for identity in all aligned species. The position of the candidate DNA sequence variant is outlined by a dashed black line and is highlighted in the multi-species alignment by a red frame. (TIF) References 1. Felius M (1995) Cattle breeds : an encyclopedia. Doetinchem: Misset. 799p. p. 2. Capitan A, Grohs C, Weiss B, Rossignol MN, Reverse P, et al. (2011) A newly described bovine type 2 scurs syndrome segregates with a frame-shift mutation in TWIST1. PLoS One 6: e22242. 3. Dove WF (1935) The physiology of horn growth: a study of the morphogenesis, the interaction of tissues and the evolutionary processes of a Mendelian recessive character by means of transplantation of tissues. J Exp Zool 69: 347–405. Allelic Heterogeneity of Bovine Polledness Figure S6 Across-species conservation for variants PT1909354A and P1909396D2. (a) The PhastCons conservation scores of the sequence around the two DNA sequence variants PT1909354A and P1909396D2 are shown. Both variants are outlined with black dotted lines. (b) Sequence identity is marked in the multi-species alignment by a red and black star among horn- bearing and all aligned species, respectively. (TIF) Table S1 Breed origin of case-control design, carriers and semi-random samples. Breed names, abbreviations, geographic origin and numbers of genotyped samples are listed for each group within breed. The four groups are homozygous polled cases (PP), heterozygous polled carriers (Pp), horned controls (pp) and the semi-random sample (R) adjusted by the diversity panel. (PDF) Table S2 Specification of the samples origin. Name of the samples supplier, affiliation, sampled breed(s), sampled tissue and primary reason for tissue sampling. This table is part of Acknowledgments and Ethics statement. (PDF) Acknowledgments We are grateful for the support of the respective numerous cattle breeders and cattle breeding associations and insemination centres that generously provided the most samples free of charge. In particular, we thank K.H. Go¨pel (Go¨pel Genetik GmbH, Germany), Dr. G. Ro¨hrmoser (Arbeitsge- meinschaft Su¨ ddt. Rinderzucht-u. Besamungsorganisationen e.V., Mu¨nchen, Germany), Dr. A. Scholz (Lehr- und Versuchsgut, Obers- chleißheim, Germany), Dr. J. Eder (Hochschule Weihenstephan-Triesdorf, Triesdorf, Germany), Dr. A. Medugorac (Institute for Animal Breeding, TU Munich), Dr. J. Ramljak (Department of Animal Science, University of Zagreb, Croatia), Dr. A. Ivankovic (Department of Animal Science, University of Zagreb, Croatia), Dr. K. Kume (NC of FAnGR, Tirana, Albania), Dr. H.P. Gru¨ nenfelder (SAVE Foundation, St. Gallen, Switzerland), Dr. V. Cadavez (Departament of Animal Science, CIMO, Braganca, Portugal), Dr. E.D. Hegemann (Tierarzt Praxis Hegemann, Soest, Germany), Dr. J. Potthast (Rinder-Union West eG, Mu¨nster, Germany), Dr. B. Weber (Masterrind GmbH, Verden, Germany), Mrs. B. Brentrup (WWS Germany GmbH, Altenberge, Germany), Mr. M. Schricker (Fleischrinderverband Bayern, Ansbach, Germany), Mr. H. Zuchtriegel (Zweckverband fu¨ r ku¨ nstliche Besamung der Haustiere, Greifenberg, Germany), Mr. E. Rosenberger (The Bavarian Gene Reserves, LfL, Grub, Germany), Mr. L. Eule (AgrarCenter Erzgebirge GmbH, Germany), Mr. N. Hartmann (Klo¨ck Hartmann GbR, Bidingen, Germany), Mr. M. Wild (Family Wild, Prittriching, Germany), Mr. S. Rist (AHG Kempten, Germany), Mr. M. Kelz (Rinderbesamungsgenos- senschaft Memmingen, Germany) and Mr. P. Lobet (Belgian Blue Group, Ciney). We thank Dr. C.E. Veit (Chair of Animal Genetics and Husbandry, LMU, Munich, Germany) for comments on English grammar. Detailed specification of the samples origin is given in the Supplementary Table S2. Author Contributions Conceived and designed the experiments: IM SR MF HB. Performed the experiments: IM SK DS. Analyzed the data: IM SR SK AG DS IR. Contributed reagents/materials/analysis tools: KHG HB MF. Wrote the paper: IM SR SK DS IR. 4. Graf B, Senn M (1999) Behavioural and physiological responses of calves to dehorning by heat cauterization with or without local anaesthesia. Applied Animal Behaviour Science 62: 153–171. 5. Ramljak J, Ivankovic A, Veit-Kensch CE, Forster M, Medugorac I (2011) Analysis of genetic and cultural conservation value of three indigenous Croatian cattle breeds in a local and global context. J Anim Breed Genet 128: 73–84. PLoS ONE | www.plosone.org 10 June 2012 | Volume 7 | Issue 6 | e39477