Chromosome Research

, 19:685 | Cite as

Anchoring the dog to its relatives reveals new evolutionary breakpoints across 11 species of the Canidae and provides new clues for the role of B chromosomes

  • Shannon E. Duke Becker
  • Rachael Thomas
  • Vladimir A. Trifonov
  • Robert K. Wayne
  • Alexander S. Graphodatsky
  • Matthew Breen


The emergence of genome-integrated molecular cytogenetic resources allows for comprehensive comparative analysis of gross karyotype architecture across related species. The identification of evolutionarily conserved chromosome segment (ECCS) boundaries provides deeper insight into the process of chromosome evolution associated with speciation. We evaluated the genome-wide distribution and relative orientation of ECCSs in three wild canid species with diverse karyotypes (red fox, Chinese raccoon dog, and gray fox). Chromosome-specific panels of dog genome-integrated bacterial artificial chromosome (BAC) clones spaced at ∼10-Mb intervals were used in fluorescence in situ hybridization analysis to construct integrated physical genome maps of these three species. Conserved evolutionary breakpoint regions (EBRs) shared between their karyotypes were refined across these and eight additional wild canid species using targeted BAC panels spaced at ∼1-Mb intervals. Our findings suggest that the EBRs associated with speciation in the Canidae are compatible with recent phylogenetic groupings and provide evidence that these breakpoints are also recurrently associated with spontaneous canine cancers. We identified several regions of domestic dog sequence that share homology with canid B chromosomes, including additional cancer-associated genes, suggesting that these supernumerary elements may represent more than inert passengers within the cell. We propose that the complex karyotype rearrangements associated with speciation of the Canidae reflect unstable chromosome regions described by the fragile breakage model.


B chromosomes fragile breakage model breakpoint reuse theory fluorescence in situ hybridization phylogenetic Canidae 



Bacterial artificial chromosome


Basic local alignment search tool


Chrysocyon brachyurus (maned wolf)


Canis familiaris (domestic dog)


Children’s Hospital Oakland Research Institute


Cellular homolog for feline sarcoma viral oncogene vKIT


Cerdocyon thous (crab-eating fox)


Deoxyribonucleic acid


Dusicyon vetulus (hoarey fox)


Evolutionary breakpoint region


Evolutionarily conserved chromosomal segment


Fragile breakage model


Fluorescence in situ hybridization


Fennecus zerda (fennec fox)


Leucine-rich repeats and immunoglobulin-like domain protein 1


Nyctereutes procynoides procynoides (Chinese raccoon dog)


Nyctereutes procynoides viverrinus (Japanese raccoon dog)


Otocyon megalotis (bat-eared fox)


Rearranged during transfection


South American


Speothus venaticus (bush dog)


Urocyon cinereogenteus (gray fox)


Vulpes macrotis (kit fox)


Vulpes vulpes (red fox)



This study was supported by a grant from the Morris Animal Foundation awarded to MB (D08ZO-022). SEDB was funded in part by the Comparative Biomedical Sciences Graduate Program at NCSU. RW was supported by funds from the National Science Foundation (DEB0614585), and ASG and VAT were supported by funds from the Program on Molecular and Cellular Biology (MCB) and Russian Foundation of Basic Research (RFBR).

Supplementary material

10577_2011_9233_Fig12_ESM.jpg (2 mb)
Supplemental Fig. 1

a Thirteen BAC probes spaced ∼10 Mb apart along the length of CFA1 were hybridized together onto CFA chromosome spreads. The location and orientation of the panel represents the CFA1 ECCS. b The CFA1 probe panel was hybridized to VVU5 and VVU1, revealing a breakpoint between probes representing CFA1;22.3 Mb (yellow) and CFA1;32. 3Mb (purple). Regions of CFA1 ECCSs on either side of the breakpoint are indicated with a suffix (i.e., CFA1a and CFA1b). Through application of the multicolor labeling strategy, the location and orientation of each CFA1 ECCS was evident (inset). Scale bar, 10 μm (JPEG 2026 kb)

10577_2011_9233_MOESM1_ESM.eps (6.2 mb)
High Resolution Image (EPS 6343 kb)
10577_2011_9233_MOESM2_ESM.xls (47 kb)
Supplemental Table 1–4 CFA regions corresponding to ECCSs are listed with the corresponding regions of red fox (VVU), Chinese raccoon dog (NPRp), and gray fox (UCI) indicated. Table 1 is sorted by dog region, Table 2 by red fox chromosome locations, Table 3 by Chinese raccoon dog chromosome locations, and Table 4 by gray fox chromosome locations. Regions with additional hybridization to B chromosomes of red fox and Chinese raccoon dog are noted with asterisks. While we used the nomenclature of Wayne et al. (1987a) for gray fox chromosomes, the final column lists the nomenclature used in Graphodatsky et al. (2008). The gray fox cells described in Graphodatsky et al. (2008) contain a translocation relative to the cells described in this text. The regions involved (CFA7, 28, 37 ECCSs on UCI12, 15) are noted with double asterisks here, shown in Fig. 4 and discussed in the text (XLS 47 kb)


  1. Alekseyev MA (2008) Multi-break rearrangements and breakpoint re-uses: from circular to linear genomes. J Comput Biol 15(8):1117–1131PubMedCrossRefGoogle Scholar
  2. Alekseyev MA, Pevzner PA (2007) Are there rearrangement hotspots in the human genome? PLoS Comp Biol 3(11):e209CrossRefGoogle Scholar
  3. Alekseyev M, Pevzner P (2010) Comparative genomics reveals birth and death of fragile regions in mammalian evolution. Genome Biol 11(11):R117–R117PubMedCrossRefGoogle Scholar
  4. Angstadt A, Motsinger-Reif A, Thomas R et al. (2011) Characterization of canine osteosarcoma by array comparative genomic hybridization and RT-qPCR: signatures of genomic imbalance in canine osteosarcoma parallel the human counterpart. Genes, Chromosomes, Cancer. doi: 10.1002/gcc.20908
  5. Bailey JA, Baertsch R, Kent WJ, Haussler D, Eichler EE (2004) Hotspots of mammalian chromosomal evolution. Genome Biol 5(4):R23PubMedCrossRefGoogle Scholar
  6. Bardeleben C, Moore RL, Wayne RK (2005) A molecular phylogeny of the Canidae based on six nuclear loci. Mol Phylogenet Evol 37(3):815–831PubMedCrossRefGoogle Scholar
  7. Breen M, Bullerdiek J, Langford CF (1999) The DAPI banded karyotype of the domestic dog (Canis familiaris) generated using chromosome-specific paint probes. Chromosome Res 7(5):401–406PubMedCrossRefGoogle Scholar
  8. Breen M, Hitte C, Lorentzen TD et al (2004) An integrated 4249 marker FISH/RH map of the canine genome. BMC Genomics 5(1):65PubMedCrossRefGoogle Scholar
  9. Camacho JP, Sharbel TF, Beukeboom LW (2000) B-chromosome evolution. Philos Trans R Soc Lond, B, Biol Sci 355(1394):163–178PubMedCrossRefGoogle Scholar
  10. Chemitiganti S, Verma RS, Silver RT, Coleman M, Dosik H (1985) Unusual translocations involving chromosomes 12;22 and 9;12 in a case of chronic myelogenous leukemia. Cancer Genet Cytogenet 14(1–2):61–65PubMedCrossRefGoogle Scholar
  11. Chen-Liu LW, Huang BC, Scalzi JM et al (1995) Selection of hybrids by affinity capture (SHAC): a method for the generation of cDNAs enriched in sequences from a specific chromosome region. Genomics 30(2):388–392PubMedCrossRefGoogle Scholar
  12. Derrien T, Andre C, Galibert F, Hitte C (2007) AutoGRAPH: an interactive web server for automating and visualizing comparative genome maps. Bioinformatics 23:498–499PubMedCrossRefGoogle Scholar
  13. Graphodatsky AS, Yang F, O'Brien PC et al (2000) A comparative chromosome map of the Arctic fox, red fox and dog defined by chromosome painting and high resolution G-banding. Chromosome Res 8(3):253–263PubMedCrossRefGoogle Scholar
  14. Graphodatsky AS, Kukekova AV, Yudkin DV et al (2005) The proto-oncogene C-KIT maps to canid B-chromosomes. Chromosome Res 13(2):113–122PubMedCrossRefGoogle Scholar
  15. Graphodatsky AS, Perelman PL, Sokolovskaya N (2008) Phylogenomics of the dog and fox family (Canidae, Carnivora) revealed by chromosome painting. Chromosome Res 16:129–146PubMedCrossRefGoogle Scholar
  16. Grzes M, Nowacka-Woszuk J, Szczerbal I, Czerwinska J, Gracz J, Switonski M (2009) A Comparison of coding sequence and cytogenetic localization of the myostatin gene in the dog, red fox, Arctic fox and Chinese raccoon dog. Cytogenet Genome Res 126:173–179PubMedCrossRefGoogle Scholar
  17. Haig D (1999) A brief history of human autosomes. Philos Trans R Soc Lond, B, Biol Sci 354(1388):1447–1470PubMedCrossRefGoogle Scholar
  18. Larkin DM, Pape G, Donthu R, Auvil L, Welge M, Lewin HA (2009) Breakpoint regions and homologous synteny blocks in chromosomes have different evolutionary histories. Genome Res 19(5):770–777PubMedCrossRefGoogle Scholar
  19. Lin KW, Yan J (2008) Endings in the middle: current knowledge of interstitial telomeric sequences. Mutat Res 658(1-2):95–110PubMedCrossRefGoogle Scholar
  20. Lindblad-Toh K, Wade CM, Mikkelsen TS et al (2005) Genome sequence, comparative analysis and haplotype structure of the domestic dog. Nature 438(7069):803–819PubMedCrossRefGoogle Scholar
  21. Ljuslinder I, Golovleva I, Palmqvist R et al (2007) LRIG1 expression in colorectal cancer. Acta Oncol 46(8):1118–1122PubMedCrossRefGoogle Scholar
  22. Ljuslinder I, Golovleva I, Henriksson R, Grankvist K, Malmer B, Hedman H (2009) Co-incidental increase in gene copy number of ERBB2 and LRIG1 in breast cancer. Breast Cancer Res 11(3):403PubMedCrossRefGoogle Scholar
  23. Mäkinen A (1985) The standard karyotype of the silver fox (Vulpes fulvus Desm.). Committee for the Standard Karyotype of Vulpes fulvus Desm. Hereditas 103(2):171–176PubMedCrossRefGoogle Scholar
  24. Mäkinen A, Kuokkanen MT, Valtonen M (1986) A chromosome-banding study in the Finnish and the Japanese raccoon dog. Hereditas 105(1):97–105PubMedCrossRefGoogle Scholar
  25. Marshall OJ, Chueh AC, Wong LH, Choo KHA (2008) Neocentromeres: new insights into centromere structure, disease development, and karyotype evolution. Am J Hum Genet 82(2):261–282PubMedCrossRefGoogle Scholar
  26. Mrózek K, Karakousis CP, Perez-Mesa C, Bloomfield CD (1993) Translocation t(12;22)(q13;q12.2-12.3) in a clear cell sarcoma of tendons and aponeuroses. Genes, Chromosom Cancer 6(4):249–252CrossRefGoogle Scholar
  27. Murphy WJ, Larkin DM, Everts-van der Wind A et al (2005) Dynamics of mammalian chromosome evolution inferred from multispecies comparative maps. Science 309(5734):613–617PubMedCrossRefGoogle Scholar
  28. Nash WG, Menninger JC, Wienberg J, Padilla-Nash HM, O'Brien SJ (2001) The pattern of phylogenomic evolution of the Canidae. Cytogenet Cell Genet 95(3–4):210–224PubMedCrossRefGoogle Scholar
  29. Nie W, Wang J, Perelman P, Graphodatsky AS, Yang F (2003) Comparative chromosome painting defines the karyotypic relationships among the domestic dog, Chinese raccoon dog and Japanese raccoon dog. Chromosome Res 11(8):735–740PubMedCrossRefGoogle Scholar
  30. Ohno S (1973) Ancient linkage groups and frozen accidents. Nature 244(5414):259–262CrossRefGoogle Scholar
  31. Peng Q, Pevzner PA, Tesler G (2006) The fragile breakage versus random breakage models of chromosome evolution. PLoS Comp Biol 2(2):e14CrossRefGoogle Scholar
  32. Pevzner P, Tesler G (2003) Human and mouse genomic sequences reveal extensive breakpoint reuse in mammalian evolution. Proc Natl Acad Sci USA 100(13):7672–7677PubMedCrossRefGoogle Scholar
  33. Pieńkowska-Schelling A, Schelling C, Zawada M, Yang F, Bugno M, Ferguson-Smith M (2008) Cytogenetic studies and karyotype nomenclature of three wild canid species: maned wolf (Chrysocyon brachyurus), bat-eared fox (Otocyon megalotis) and fennec fox (Fennecus zerda). Cytogenet Genome Res 121(1):25–34PubMedCrossRefGoogle Scholar
  34. Piras FM, Nergadze SG, Magnani E et al (2010) Uncoupling of satellite DNA and centromeric function in the genus Equus. PLoS Genet 6(2):1–10CrossRefGoogle Scholar
  35. Ruiz-Herrera A, García F, Mora L, Egozcue J, Ponsà M, Garcia M (2005) Evolutionary conserved chromosomal segments in the human karyotype are bounded by unstable chromosome bands. Cytogenet Genome Res 108(1–3):161–174PubMedCrossRefGoogle Scholar
  36. Schröder C, Bleidorn C, Hartmann S, Tiedemann R (2009) Occurrence of Can-SINEs and intron sequence evolution supports robust phylogeny of pinniped carnivores and their terrestrial relatives. Gene 448(2):221–226PubMedCrossRefGoogle Scholar
  37. Thomas R, Duke SE, Bloom SK et al (2007) A cytogenetically characterized, genome-anchored 10-Mb BAC set and CGH array for the domestic dog. J Hered 98(5):474–484PubMedCrossRefGoogle Scholar
  38. Thomas R, Duke SE, Karlsson EK et al (2008) A genome assembly-integrated dog 1 Mb BAC microarray: a cytogenetic resource for canine cancer studies and comparative genomic analysis. Cytogenet Genome Res 122(2):110–121PubMedCrossRefGoogle Scholar
  39. Thomas R, Duke SE, Wang HJ et al (2009) 'Putting our heads together': insights into genomic conservation between human and canine intracranial tumors. J Neurooncol 94(3):333–349PubMedCrossRefGoogle Scholar
  40. Thomas R, Seiser EL, Motsinger-Reif A et al (2011) Refining tumor-associated aneuploidy through 'genomic recoding' of recurrent DNA copy number aberrations in 150 canine non-Hodgkin lymphomas. Leuk Lymphoma 52(7):1321–1335PubMedCrossRefGoogle Scholar
  41. Trifonov VA, Perelman PL, Kawada SI, Iwasa MA, Oda SI, Graphodatsky AS (2002) Complex structure of B-chromosomes in two mammalian species: Apodemus peninsulae (Rodentia) and Nyctereutes procyonoides (Carnivora). Chromosome Res 10(2):109–116PubMedCrossRefGoogle Scholar
  42. Vujosević M, Blagojević J (2004) B chromosomes in populations of mammals. Cytogenet Genome Res 106(2–4):247–256PubMedGoogle Scholar
  43. Wade CM, Giulotto E, Sigurdsson S et al (2009) Genome sequence, comparative analysis, and population genetics of the domestic horse. Science 326(5954):865–867PubMedCrossRefGoogle Scholar
  44. Ward OG, Wurster-Hill DH, Ratty FJ, Song Y (1987) Comparative cytogenetics of Chinese and Japanese raccoon dogs, Nyctereutes procyonoides. Cytogenet Cell Genet 45(3–4):177–186PubMedCrossRefGoogle Scholar
  45. Wayne RK (1993) Molecular evolution of the dog family. Trends Genet 9(6):218–224PubMedCrossRefGoogle Scholar
  46. Wayne RK, Ostrander EA (1999) Origin, genetic diversity, and genome structure of the domestic dog. Bioessays 21(3):247–257PubMedCrossRefGoogle Scholar
  47. Wayne RK, Nash WG, O'Brien SJ (1987a) Chromosomal evolution of the Canidae. I. Species with high diploid numbers. Cytogenet Cell Genet 44(2–3):123–133PubMedCrossRefGoogle Scholar
  48. Wayne RK, Nash WG, O'Brien SJ (1987b) Chromosomal evolution of the Canidae. II. Divergence from the primitive carnivore karyotype. Cytogenet Cell Genet 44(2–3):134–141PubMedCrossRefGoogle Scholar
  49. Wayne RK, Geffen E, Girman DJ, Koepfli KP, Lau LM, Marshall CR (1997) Molecular systematics of the Canidae. Syst Biol 46(4):622–653PubMedCrossRefGoogle Scholar
  50. Yang F, O'Brien PC, Milne BS et al (1999) A complete comparative chromosome map for the dog, red fox, and human and its integration with canine genetic maps. Genomics 62(2):189–202PubMedCrossRefGoogle Scholar
  51. Yang W-M, Yan Z-J, Ye Z-Q, Guo D-S (2006) LRIG1, a candidate tumour-suppressor gene in human bladder cancer cell line BIU87. BJU Int 98(4):898–902PubMedCrossRefGoogle Scholar
  52. Yudkin DV, Trifonov VA, Kukekova AV et al (2007) Mapping of KIT adjacent sequences on canid autosomes and B chromosomes. Cytogenet Genome Res 116(1–2):100–103PubMedCrossRefGoogle Scholar
  53. Zrzavy J, Ricankova V (2004) Phylogeny of recent Canidae (Mammalia, Carnivora): relative reliability and utility of morphological and molecular datasets. Zoologica Scripta 33:311–333CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Shannon E. Duke Becker
    • 1
  • Rachael Thomas
    • 1
    • 2
  • Vladimir A. Trifonov
    • 3
  • Robert K. Wayne
    • 4
  • Alexander S. Graphodatsky
    • 3
  • Matthew Breen
    • 1
    • 2
    • 5
  1. 1.Department of Molecular Biomedical Sciences, College of Veterinary MedicineNorth Carolina State UniversityRaleighUSA
  2. 2.Center for Comparative Medicine and Translational ResearchNorth Carolina State UniversityRaleighUSA
  3. 3.Department of Molecular and Cellular BiologyInstitute of Chemical Biology and Fundamental MedicineNovosibirskRussia
  4. 4.Department of Ecology and Evolutionary BiologyUniversity of California Los AngelesLos AngelesUSA
  5. 5.Cancer Genetics Program, UNC Lineberger Comprehensive Cancer CenterChapel HillUSA

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