, Volume 127, Issue 3, pp 323–340 | Cite as

High-throughput analysis of satellite DNA in the grasshopper Pyrgomorpha conica reveals abundance of homologous and heterologous higher-order repeats

  • Francisco J. Ruiz-Ruano
  • Jesús Castillo-Martínez
  • Josefa Cabrero
  • Ricardo Gómez
  • Juan Pedro M. Camacho
  • María Dolores López-LeónEmail author
Original Article


Satellite DNA (satDNA) constitutes an important fraction of repetitive DNA in eukaryotic genomes, but it is barely known in most species. The high-throughput analysis of satDNA in the grasshopper Pyrgomorpha conica revealed 87 satDNA variants grouped into 76 different families, representing 9.4% of the genome. Fluorescent in situ hybridization (FISH) analysis of the 38 most abundant satDNA families revealed four different patterns of chromosome distribution. Homology search between the 76 satDNA families showed the existence of 15 superfamilies, each including two or more families, with the most abundant superfamily representing more than 80% of all satDNA found in this species. This also revealed the presence of two types of higher-order repeats (HORs), one showing internal homologous subrepeats, as conventional HORs, and an additional type showing non-homologous internal subrepeats, the latter arising by the combination of a given satDNA family with a non-annotated sequence, or with telomeric DNA. Interestingly, the heterologous subrepeats included in these HORs showed higher divergence within the HOR than outside it, suggesting that heterologous HORs show poor homogenization, in high contrast with conventional (homologous) HORs. Finally, heterologous HORs can show high differences in divergence between their constituent subrepeats, suggesting the possibility of regional homogenization.


FISH Higher-order repeat (HOR) High-throughput sequencing Pyrgomorpha conica Satellitome 



We thank Teresa Palomeque, Pedro Lorite and Manuel Garrido for their valuable comments on the manuscript. This study was funded by grants from Spanish Plan Andaluz de Investigación (CVI-6649) and Secretaría de Estado de Investigación, Desarrollo e Innovación (CGL2015-70750-P) and was partially performed by FEDER funds. F.J. Ruiz-Ruano was supported by a Junta de Andalucía fellowship.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed

Supplementary material

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ESM 1 (PDF 1232 kb)
412_2018_666_MOESM2_ESM.pdf (16 kb)
ESM 2 (PDF 16 kb)


  1. Antonio C, González-García JM, Suja JA (1993) Pycnotic cycle of the sex chromosome in Pyrgomorpha conica (Orthoptera) and development of spermiogenesis. Genome 36:535–541. CrossRefPubMedGoogle Scholar
  2. Benson G (1999) Tandem repeats finder: a program to analyze DNA sequences. Nucleic Acids Res 27:573–580CrossRefPubMedPubMedCentralGoogle Scholar
  3. Blanchetot A (1991) Genetic variability of a satellite sequences in the dipteran Musca domestica. EXS 58:106–112PubMedGoogle Scholar
  4. Britten RJ, Kohne DE (1968) Repeated sequences in DNA. Science 161:529–540CrossRefPubMedGoogle Scholar
  5. Bonaccorsi S, Lohe A (1991) Fine mapping of satellite DNA sequences along the Y chromosomes of Drosophila melanogaster: relationship between satellite sequences and fertility factors. Genetics 129:177–189PubMedPubMedCentralGoogle Scholar
  6. Camacho JPM, Cabrero J, López-León MD, Cabral-de Mello DC, Ruiz-Ruano FJ (2015b) Grasshoppers (Orthoptera). In Sharakhov IV (ed) Protocols for cytogenetic mapping of arthropod genomes. CRC Press, pp 381-438Google Scholar
  7. Camacho JPM, Ruiz-Ruano FJ, Martín-Blázquez R, López-León MD, Cabrero J, Lorite P, Cabral-de-Mello DC, Bakkali M (2015a) A step to the gigantic genome of the desert locust: chromosome sizes and repeated DNAs. Chromosoma 124:263–275. CrossRefPubMedGoogle Scholar
  8. Charlesworth B, Sniegowsky P, Stephan W (1994) The evolutionary dynamics of repetitive DNA in eukaryotes. Nature 371:215–220. CrossRefPubMedGoogle Scholar
  9. Cohen S, Agmon N, Sobol O, Segal D (2010) Extrachromosomal circles of satellite repeats and 5S ribosomal DNA in human cells. Mob DNA 1:11. CrossRefPubMedPubMedCentralGoogle Scholar
  10. Cohen S, Agmon N, Yacobi K, Mislovati M, Segal D (2005) Evidence for rolling circle replication of tandem genes in Drosophila. Nucleic Acids Res 33:4519–4526. CrossRefPubMedPubMedCentralGoogle Scholar
  11. Cohen S, Houben A, Segal D (2008) Extrachromosomal circular DNA derived from tandemly repeated genomic sequences in plants. Plant J 53:1027–1034. CrossRefPubMedGoogle Scholar
  12. Drummond AJ, Ashton B, Cheung M, Heled J, Kearse M (2009) Geneious 4.8. Biomatters Ltd, Auckland, New ZealandGoogle Scholar
  13. Excoffier L, Lischer HE (2010) Arlequin suite ver 3.5: a new series of programs to perform population genetics analyses under Linux and Windows. Mol Ecol Resour 10:564–567. CrossRefPubMedGoogle Scholar
  14. Feliciello I, Akrap I, Ugarkovic (2015) Satellite DNA modulats gene expression in beetle Tribolium castaneum after heat stress. PLoS Genet 11:e1005466. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Feliciello I, Picariello O, Chinali G (2006) Intra-specific variability and unusual organization of the repetitive units in a satellite DNA from Rana dalmatina: molecular evidence of a new mechanism of DNA repair acting on satellite DNA. Gene 383:81–92. CrossRefPubMedGoogle Scholar
  16. Fukagawa T, Earnshaw WC (2014) The centromere: chromatin foundation for the kinetochore machinery. Dev Cell 30:496–508. CrossRefPubMedPubMedCentralGoogle Scholar
  17. García G, Ríos N, Gutiérrez V (2015) Next generation sequencing detects repetitive elements expansion in giant genomes of annual killifish genus Austrolebias (Cyprinodontiformes, Rivulidae). Genetica 143:353–360. CrossRefPubMedGoogle Scholar
  18. Garrido-Ramos MA (2017) Satellite DNA: an evolving topic. Genes 8:230. CrossRefPubMedCentralGoogle Scholar
  19. Giunta S, Funabiki H (2017) Integrity of the human centromere DNA repeats is protected by CENP-A, CENP-C, and CENP-T. Proc Natl Acad Sci U S A 114:1928–1933. CrossRefPubMedPubMedCentralGoogle Scholar
  20. Greider CW, Blackburn EH (1985) Identification of a specific telomere terminal transferase activity in tetrahymena extracts. Cell 43(2 Pt 1):405–413. CrossRefPubMedGoogle Scholar
  21. Hahn C, Bachmann L, Chevreux B (2013) Reconstructing mitochondrial genomes directly from genomic next-generation sequencing reads—a baiting and iterative mapping approach. Nucleic Acids Res 41:e129. CrossRefPubMedPubMedCentralGoogle Scholar
  22. Henikoff S (1998) Conspiracy of silence among repeated transgenes. Bioessay 20:532–535.<532::AID-BIES3>3.0.CO;2-M CrossRefGoogle Scholar
  23. Hemleben V, Torres-Ruiz R, Schmidt T, Zentgraf U (2000) Molecular cell biology: role of repetitive DNA in nuclear architecture and chromosome structure. In: Progress in botany vol 61. Springer, Germany, pp 91–117CrossRefGoogle Scholar
  24. Hsieh J, Fire A (2000) Recognition and silencing of repeated DNA. Annu Rev Genet 34:187–204. CrossRefPubMedGoogle Scholar
  25. John B (1988) The biology of heterochromatin. In: RS Verman (ed) Heterochromatin, molecular and structural aspects. Cambridge University Press, Cambridge, pp. 1–147Google Scholar
  26. Kass DH, Batzer MA (2001) Genome organization:human. ELS:1–8.
  27. Kent WJ (2002) BLAT—the BLAST-like alignment tool. Genome Res 12:656–664. CrossRefPubMedPubMedCentralGoogle Scholar
  28. Kuhn GC, Küttler H, Moreira-Filho O, Heslop-Harrison JS (2012) The 1.688 repetitive DNA of Drosophila: concerted evolution at different genomic scales and association with genes. Mol Biol Evol 29:7–11. CrossRefPubMedGoogle Scholar
  29. Langmead B, Salzberg SL (2012) Fast gapped-read alignment with bowtie 2. Nat Methods 9:357–359. CrossRefPubMedPubMedCentralGoogle Scholar
  30. Lica LM, Narayanswami S, Hamkalo BA (1986) Mousse satellite DNA, centromere structure and sister chromatid pairing. J Cell Biol 103:1045–1151CrossRefGoogle Scholar
  31. Littlewood DTJ, Olson PD (2001) Small subunit rDNA and the platyhelminthes: signal, noise, conflict and compromise. In D.T.J. Littlewood & R.A. Bray (eds) Interrelationships of the Platyhelminthes .CRC Press, UK, pp, 1–33Google Scholar
  32. López-Fernández C, Arroyo F, Fenández JL, Gosálvez J (2006) Interstitial telomeric sequence blocks in constitutive pericentromeric heterochromatin from Pyrgomorpha conica (Orthoptera) are enriched in constitutive alkali-labile sites. Mut Res 599:36–44. CrossRefGoogle Scholar
  33. López-Fernández C, Gosálvez J, Suja JA, Mezzanotte (1988) Restriction endonuclease digestion of meiotic and mitotic chromosomes in Pyrgomorpha conica (Orthoptera: Pyrgomorphidae). Genome 30:621–626. CrossRefGoogle Scholar
  34. López-Fernández C, Pradillo E, Zabal-Aguirre M, Fenández JL, de la Vega G, C, Gosálvez J (2004) Telomeric and interstitial telomeric-like DNA sequences in Orthoptera genomes. Genome 47:757–763.
  35. López-Flores I, Garrido-Ramos MA (2012) The repetitive DNA content of eukaryotic genomes. In: Garrido-Ramos MA (ed) Repetitive DNA. Genome Dyn, 7, Karger, Basel, pp 1-28. doi:
  36. Macas J, Kejnovský E, Neumann P, Novák P, Koblížková A, Vyskot B (2011) Next generation sequencing-based analysis of repetitive DNA in the model dioceous plant Silene latifolia. PLoS One 6:e27335. CrossRefPubMedPubMedCentralGoogle Scholar
  37. Macas J, Neumann P, Navrátilová A (2007) Repetitive DNA in the pea (Pisum sativum L.) genome: comprehensive characterization using 454 sequencing and comparison to soybean and Medicago truncatula. BMC Genomics 8:427. CrossRefPubMedPubMedCentralGoogle Scholar
  38. Meštrović N, Mravinac B, Pavlek M, Vojvoda-Zeljko T, Šatović E, Plohl M (2015) Structural and functional liaisons between transposable elements and satellite DNAs. Chromosom Res 23:583–596. CrossRefGoogle Scholar
  39. Meyne J, Hirai H, Imai HT (1995) FISH analysis of the telomere sequences of bulldog ants (Myrmecia: Formidae). Chromosoma 104:14–18. PubMedCrossRefGoogle Scholar
  40. Miga KH (2015) Completing the human genome: the progress and challenge of satellite DNA assambly. Chromosom Res 23:421–426. CrossRefGoogle Scholar
  41. Mravinac B, Plohl M (2007) Satellite DNA junctions identify the potential origin of new repetitive elements in the beetle Tribolium madens. Gene 394:45–52. CrossRefPubMedGoogle Scholar
  42. Navrátilová A, Koblížková A, Macas J (2008) Survey of extrachromosomal circular DNA derived from plant satellite repeats. BMC Plant Biol 8:90. CrossRefPubMedPubMedCentralGoogle Scholar
  43. Novák P, Hribová E, Neumann P, Koblízková A, Dolezel J, Macas J (2014) Genome-wide analysis of repeat diversity across the family musaceae. PLoS One 9(6):e98918 doi:.1371/journal.pone.0098918CrossRefPubMedPubMedCentralGoogle Scholar
  44. Novák P, Neumann P, Pech J, Steinhaisl J, Macas J (2013) RepeatExplorer: a galaxy-based web server for genome-wide characterization of eukaryotic repetitive elements from next-generation sequence reads. Bioinformatics 29:792–793. CrossRefPubMedGoogle Scholar
  45. Palomeque T, Lorite P (2008) Satelite DNA in insects: a review. Heredity 100:564–573. CrossRefPubMedGoogle Scholar
  46. Plohl M, Luchetti A, Meštrović N, Mantovani B (2008) Satellite DNAs between selfishness and functionality: structure, genomics and evolution of tandem repeats in centromeric (hetero)chromatin. Gene 409:72–82. CrossRefPubMedGoogle Scholar
  47. Plohl M, Meštrović N, Bruvo B, Ugarkovic (1998) Similarity of structural features and evolution of satellite DNAs from Palorus subdepressus (Coleoptera) and related species. J Mol Evol 46:234–239Google Scholar
  48. Plohl M, Meštrović N, Mravinac B (2012) Satellite DNA evolution. In: Garrido-Ramos MA (ed) Repetitive DNA, vol 7. Karger, Basel, pp 126–152. CrossRefGoogle Scholar
  49. Plohl M, Meštrović N, Mravinac B (2014) Centromere identity from the DNA point of view. Chromosoma 123:313–325. CrossRefPubMedPubMedCentralGoogle Scholar
  50. Rossi MS, Reig OA, Zorzópulos J (1990) Evidence for rolling-circle replication in a major satellite DNA from the South American rodents of the genus Ctenomys. Mol Biol Evol 7:340–350PubMedGoogle Scholar
  51. Ruiz-Estévez M, Cabrero J, Camacho JPM, López-León MD (2014) B chromosomes in the grasshopper Eyprepocnemis plorans are present in all body parts analyzed and show extensive variation for rDNA copy number. Cytogenet Genome Res 143:268–274. CrossRefPubMedGoogle Scholar
  52. Ruiz-Ruano FJ, Cabrero J, López-León MD, Camacho JPM (2017) Satellite DNA content illuminates the ancestry of a supernumerary (B) chromosome. Chromosoma 126:487–500. CrossRefPubMedGoogle Scholar
  53. Ruiz-Ruano FJ, Cuadrado Á, Montiel EE, Camacho JPM, López-León MD (2015) Next generation sequencing and FISH reveal uneven and nonrandom microsatellite distribution in two grasshopper genomes. Chromosoma 124:221–234. CrossRefPubMedGoogle Scholar
  54. Ruiz-Ruano FJ, López-León MD, Cabrero J, Camacho JPM (2016) High-throughput analysis of the satellitome illuminates satellite DNA evolution. Sci Rep 6:28333. CrossRefPubMedPubMedCentralGoogle Scholar
  55. Ruiz-Ruano FJ, Ruiz-Estévez M, Rodríguez-Pérez J, López-Pino JL, Cabrero J, Camacho JPM (2011) DNA amount of X and B chromosomes in the grasshoppers Eyprepocnemis plorans and Locusta migratoria. Cytogenet Genome Res 134:120–126. CrossRefPubMedGoogle Scholar
  56. Santos JL, Arana P, Giráldez R (1983) Chromosome banding patterns in Spanish Acridoidea. Genetica 61:65–74. CrossRefGoogle Scholar
  57. Schmieder R, Edwards R (2011) Fast identification and removal of sequence contamination from genomic and metagenomic datasets. PLoS One 6:e17288. CrossRefPubMedPubMedCentralGoogle Scholar
  58. Schueler MG, Higgins AW, Rudd MK, Gustashaw K, Willard HF (2001) Genomic and genetic definition of a functional human centromere. Science 294:109–115. CrossRefPubMedGoogle Scholar
  59. Smit AFA, Hubley R, Green P (2013) RepeatMasker Open-4.0. <>
  60. Song H, Amédégnato C, Cigliano MM, Desutter-Grandcolas L, Heads SW, Huang Y, Otte D, Whiting MF (2015) 300 million years of diversification: elucidating the patterns of orthopteran evolution based on comprehensive taxon and gene sampling. Cladistics 31:621–651. CrossRefGoogle Scholar
  61. Stephan W, Cho S (1994) Possible role of natural selection in the formation of tandem- repetitive noncoding DNA. Genetics 136:333–341PubMedPubMedCentralGoogle Scholar
  62. Suja JA, Antonio C, González-García JM, Rufas JS (1993) Supernumeary heterochromatin segments associated with nucleolar chromosomes of Pyrgomorpha conica (Orthoptera) contain methylated rDNA sequences. Chromosoma 102:491–499. CrossRefPubMedGoogle Scholar
  63. Sujiwattanarat P, Thapana W, Srikulnath K, Hirai Y, Hirai H, Koga A (2015) Higher-order repeat structure in alpha satellite DNA occurs in New World monkeys and is not confined to hominoids. Sci Rep 5:10315. CrossRefPubMedPubMedCentralGoogle Scholar
  64. Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739. CrossRefPubMedPubMedCentralGoogle Scholar
  65. Thorvaldsdóttir H, Robinson JT, Mesirov JP (2013) Integrative genomics viewer (IGV): high-performance genomics data visualization and exploration. Brief Bioinform 14:178–192. CrossRefPubMedGoogle Scholar
  66. Ugarković D, Plohl M (2002) Variation in satellite DNA profiles—causes and effects. EMBO J 21:5955–5959. CrossRefPubMedGoogle Scholar
  67. Untergrasser A, Cutcutache I, Koressaar T, Ye J, Faircloth BC, Remm M, Rozen SG (2012) Primer3—new capabilities and interfaces. Nucleic Acids Res 40:e115. CrossRefGoogle Scholar
  68. Van Dijk EL, Auger H, Jaszczyszyn Y, Thermes C (2014) Ten years of next generation sequencing technology. Trends Genet 30:418–426. CrossRefPubMedGoogle Scholar
  69. Walsh JB (1987) Persistence of tandem arrays: implications for satellite and simple-sequence DNAs. Genetics 115:553–567PubMedPubMedCentralGoogle Scholar
  70. Warburton PE, Willard HF (1990) Genomic analysis of sequence variation in tandemly repeated DNA. Evidence for localized homogeneous sequence domains within arrays of alpha-satellite DNA. J Mol Biol 216:3–16CrossRefPubMedGoogle Scholar
  71. Willard HF, Waye JS (1987) Chromosome-specific subsets of human alpha satellite DNA: analysis of sequence divergence within and between chromosomal subsets and evidence for an ancestral pentameric repeat. J Mol Evol 25:207–214. CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Francisco J. Ruiz-Ruano
    • 1
  • Jesús Castillo-Martínez
    • 1
    • 3
  • Josefa Cabrero
    • 1
  • Ricardo Gómez
    • 2
  • Juan Pedro M. Camacho
    • 1
  • María Dolores López-León
    • 1
    Email author
  1. 1.Departamento de Genética. Facultad de CienciasUniversidad de GranadaGranadaSpain
  2. 2.Departamento de Ciencia y Tecnología Agroforestal, E.T.S. de Ingenieros AgrónomosUniversidad de Castilla La ManchaAlbaceteSpain
  3. 3.Facultad de MedicinaUniversidad Católica de ValenciaValenciaSpain

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