, Volume 95, Issue 4, pp 271–284 | Cite as

Chromosome banding in Amphibia

XI. Constitutive heterochromatin, nucleolus organizers, 18S + 28S and 5S ribosomal RNA genes in Ascaphidae, Pipidae, Discoglossidae and Pelobatidae
  • M. Schmid
  • L. Vitelli
  • R. Batistoni


The karyotypes of 14 species of Anura from 9 genera of the suborders Amphicoela, Aglossa, Opisthocoela and Anomocoela were analysed with various banding techniques and conventional cytogenetic methods. The 18S + 28S and 5S ribosomal RNA genes were localized by means of in situ hybridization. No Q-, R- and G-banding patterns in the euchromatic segments of the metaphase chromosomes could be demonstrated in any of the species; this does not seem to be caused by a higher degree of spiralization of the amphibian chromosomes, but by the special DNA organization in these organisms. In most karyotypes, constitutive heterochromatin is present at centromeres, telomeres and nucleolus organizer regions (NORs), but rarely in interstitial positions. The heterochromatic regions are either quinacrine positive and mithramycin negative or vice versa. All species examined possess only one homologous pair of NORs; these display the brightest mithramycin fluorescence in the karyotypes. Many specimens exhibited unequal labelling of the two NORs both after silver and mithramycin staining as well as after in situ hybridization with 3H-18S + 28S rRNA. In four species, between one and six chromosome pairs with homologous 5S rRNA sites could be identified. The 5S rRNA genes and the 18S + 28S rRNA genes are closely linked in two species. In the male meiosis of the Amphicoela and Opisthocoela, there are intersitial, subterminal and terminal chiasmata in the bivalents, whereas only terminal chiasmata are observed in the bivalents of the Aglossa and Anomocoela. No heteromorphic sex-specific chromosomes could be demonstrated in any of the species. The differential staining techniques revealed that the chromosomal structure in these four suborders is largely the same as in the highly evolved anuran suborders Procoela and Diplasiocoela.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Bachmann K (1972) Genome size in mammals. Chromosoma 37:85–93Google Scholar
  2. Bailly S (1976) Localisation et signification des zones Q observées sur les chromosomes mitotiques de l'amphibien Pleurodeles waltlii Michah. après coloration par la moutarde de quinacrine. Chromosoma 54:61–68Google Scholar
  3. Barr HJ, Esper H (1963) Nucleolar size in cells of Xenopus laevis in relation to nucleolar competition. Exp Cell Res 31:211–214Google Scholar
  4. Barsacchi-Pilone G, Nardi I, Batistoni R, Andronico F, Beccari E (1974) Chromosome location of the genes for 28S, 18S and 5S ribosomal RNA in Triturus marmoratus (Amphibia Urodela). Chromosoma 49:135–153Google Scholar
  5. Barsacchi-Pilone G, Nardi I, Andronico F, Batistoni R, Durante M (1977) Chromosomal location of the ribosomal RNA genes in Triturus vulgaris meridionalis (Amphibia, Urodela). I. Localization of the DNA sequences complementary to 5S rRNA on mitotic and lampbrush chromosomes. Chromosoma 63:127–134Google Scholar
  6. Bernardi G, Olofsson B, Filipski J, Zerial M, Salinas J, Cuny G, Meunier-Rotival M, Rodier M (1985) The mosaic genome of warm-blooded vertebrates. Science 228:953–958Google Scholar
  7. Birnstiel ML, Wallace H, Sirlin JL, Fischberg M (1966) Localization of the ribosomal DNA complements in the nucleolus organizer region of Xenopus laevis. In: Vincent WS, Miller OL (eds) International symposium on the nucleolus. Its structure and function. NCI monograph 23, National Cancer Institute, Bethesda, pp 431–448Google Scholar
  8. Birstein VJ (1982) Structural characteristics of genome organization in amphibians: differential staining of chromosomes and DNA structure. J Mol Evol 18:73–91Google Scholar
  9. Comings DE (1975) Mechanisms of chromosome banding. VIII. Hoechst 33 258-DNA interactions. Chromosoma 52:229–243Google Scholar
  10. De Lucchini S, Vitelli L, Batistoni R (1981) Bandeggio cromosomico in alcune specie di anfibi anuri. I. “C-banding”. Atti Soc Tosc Sci Nat Mem, Serie B, 88:93–101Google Scholar
  11. Elsdale TR, Fischberg M, Smith SA (1958) A mutation that reduces nucleolar number in Xenopus laevis. Exp Cell Res 14:642–643Google Scholar
  12. Estes R (1970) New fossil pelobatid frogs and a review of the genus Eopelobates. Bull Mus Comp Zool 139:337–340Google Scholar
  13. Estes R, Reig OA (1973) The early fossil record of frogs. A review of the evidence. In: Vial JL (ed) Evolutionary biology of the anurans. Univ Missouri Press, Columbia, Missouri, pp 11–63Google Scholar
  14. Hecht MK (1963) A reevaluation of the history of the frogs. II. Syst Zool 12:20–35Google Scholar
  15. Hennen S, Mizuno S, Macgregor HC (1975) In situ hybridization of ribosomal DNA labeled with 125iodine to metaphase and lampbrush chromosomes from newts. Chromosoma 50:349–369Google Scholar
  16. Holmquist G (1987) DNA sequences in G-bands and R-bands. In: Adolph KW (ed) Chromosome and chromatin structure. CRC Press, Boca Raton, Florida, in pressGoogle Scholar
  17. Kahn J (1962) The nucleolar organizer in the mitotic chromosome complement in Xenopus laevis. Q J Microsc Sci 103:407–409Google Scholar
  18. Kezer J, Macgregor HC (1973) The nucleolar organizer of Plethodon cinereus cinereus (Green). II. The lampbrush nucleolar organizer. Chromosoma 42:427–444Google Scholar
  19. Kuro-o M, Ikebe C, Kohno S (1986) Cytogenetic studies of Hynobiidae (Urodela). IV. DNA replication bands (R-banding) in the genus Hynobius and the banding karyotype of Hynobius nigrescens Stejneger. Cytogenet Cell Genet 43:14–18Google Scholar
  20. Lubs H, Hostetter T, Ewing L (1972) Paris Conference: Standardization in human cytogenetics. Birth defects. Original article series, vol 8, no 7. The National Foundation, New YorkGoogle Scholar
  21. Macgregor HC, Kezer J (1973) The nucleolar organizer of Plethodon cinereus cinereus (Green). I. Location of the nucleolar organizer by in situ nucleic acid hybridization. Chromosoma 42:415–426Google Scholar
  22. Macgregor HC, Mizuno S (1976) In situ hybridization of nicktranslated 3H-ribosomal DNA to chromosomes from salamanders. Chromosoma 54:15–25Google Scholar
  23. Macgregor HC, Vlad M, Barnett L (1977) An investigation of some problems concerning nucleolus organizers in salamanders. Chromosoma 59:283–299Google Scholar
  24. Mayol J, Alcover JA (1981) Survival of Baleaphryne Sanchiz and Adrover, 1979 (Amphibia: Anura: Discoglossidae) on Mallorca. Amphibia-Reptilia 3/4:343–345Google Scholar
  25. Mayol J, Alcover JA, Alomar G, Pomar G, Jurado J, Jaume D (1980) Supervivència de Baleaphryne (Amphibia: Anura: Discoglossidae) a les muntanyes de Mallorca. Nota preliminar. Butll Inst Cat Hist Nat 45 (Sec Zool, 3):115–119Google Scholar
  26. Miller L, Brown DD (1969) Variation in the activity of nucleolar organizers and their ribosomal gene content. Chromosoma 28:430–444Google Scholar
  27. Miller L, Knowland J (1970) Reduction of ribosomal RNA synthesis and ribosomal RNA genes in a mutant of Xenopus laevis which organizes only a partial nucleolus. II. The number of ribosomal RNA genes in animals of different nucleolar types. J Mol Biol 53:329–338Google Scholar
  28. Morescalchi A (1973) Amphibia. In: Chiarelli AB, Capanna E (eds) Cytotaxonomy and vertebrate evolution. Academic Press, London New York, pp 233–348Google Scholar
  29. Morescalchi A (1980) Evolution and karyology of the amphibians. Boll Zool 47 (Suppl):113–126Google Scholar
  30. Nardi I, Barsacchi-Pilone G, Batistoni R, Andronico F (1977) Chromosome location of the ribosomal RNA genes in Triturus vulgaris meridionalis (Amphibia, Urodela). II. Intraspecific variability in number and position of the chromosome loci for 18S + 28S ribosomal RNA. Chromosoma 64:67–84Google Scholar
  31. Nardi I, De Lucchini S, Barsacchi-Pilone G, Andronico F (1978) Chromosome location of the ribosomal RNA genes in Triturus vulgaris meridionalis (Amphibia, Urodela). IV. Comparison between in situ hybridization with 3H 18S + 28S rRNA and ASSAT staining. Chromosoma 70:91–99Google Scholar
  32. Olmo E, Morescalchi A (1978) Genome and cell sizes in frogs: a comparison with salamanders. Experientia 34:44–46Google Scholar
  33. Olmo E, Morescalchi A, Stingo V, Odierna G (1982) Genome characteristics and the systematics of the Discoglossidae (Amphibia, Salientia). Monit Zool Ital 16:283–299Google Scholar
  34. Olofsson B, Bernardi G (1983) Organization of nucleotide sequences in the chicken genome. Eur J Biochem 130:241–245Google Scholar
  35. Pardue ML (1973) Localization of repeated DNA sequences in Xenopus chromosomes. Cold Spring Harbor Symp Quant Biol 38:475–482Google Scholar
  36. Pardue ML, Gall JG (1975) Nucleic acid hybridization to the DNA of cytological preparations. Methods Cell Biol 10:1–16Google Scholar
  37. Pardue ML, Brown DD, Birnstiel ML (1973) Location of the genes for 5S ribosomal RNA in Xenopus laevis. Chromosoma 42:191–203Google Scholar
  38. Ragghianti M, Bucci Innocenti S, Mancino G (1973) Bandeggiatura indotta da “C-, G-e Q-staining methods” e pattern di replicazione dei cromosomi di Triturus. Rend Accad Naz Lincei 55:764–770Google Scholar
  39. Sanchíz FB, Adrover R (1979) Anfibios fósiles del Pleistoceno de Mallorca. Doñana, Acta Vertebrata 4:5–25Google Scholar
  40. Schempp W, Schmid M (1981) Chromosome banding in Amphibia. VI. BrdU-replication patterns in Anura and demonstration of XX/XY sex chromosomes in Rana esculenta. Chromosoma 83:697–710Google Scholar
  41. Schmid M (1978a) Chromosome banding in Amphibia. I. Constitutive heterochromatin and nucleolus organizer regions in Bufo and Hyla. Chromosoma 66:361–388Google Scholar
  42. Schmid M (1978b) Chromosome banding in Amphibia. II. Constitutive heterochromatin and nucleolus organizer regions in Ranidae, Microhylidae and Rhacophoridae. Chromosoma 68:131–148Google Scholar
  43. Schmid M (1980a) Chromosome evolution in Amphibia. In: Müller H (ed) Cytogenetics of vertebrates. Birkhäuser, Basel Boston Stuttgart, pp 4–27Google Scholar
  44. Schmid M (1980b) Chromosome banding in Amphibia. IV. Differentiation of GC-and AT-rich chromosome regions in Anura. Chromosoma 77:83–103Google Scholar
  45. Schmid M (1980c) Chromosome banding in Amphibia. V. Highly differentiated ZW/ZZ sex chromosomes and exceptional genome size in Pyxicephalus adspersus (Anura, Ranidae). Chromosoma 80:69–96Google Scholar
  46. Schmid M (1982) Chromosome banding in Amphibia. VII. Analysis of the structure and variability of NORs in Anura. Chromosoma 87:327–344Google Scholar
  47. Schmid M, Olert J, Klett C (1979) Chromosome banding in Amphibia. III. Sex chromosomes in Triturus. Chromosoma 71:29–55Google Scholar
  48. Schmid M, Löser C, Schmidtke J, Engel W (1982) Evolutionary conservation of a common pattern of activity of nucleolus organizers during spermatogenesis in vertebrates. Chromosoma 86:149–179Google Scholar
  49. Schnedl W, Breitenbach M, Mikelsaar A-V, Stranzinger G (1977) Mithramycin and DIPI: a pair of fluorochromes specific for GC-and AT-rich DNA respectively. Hum Genet 36:299–305Google Scholar
  50. Schweizer D (1976) Reverse fluorescent chromosome banding with chromomycin and DAPI. Chromosoma 58:307–324Google Scholar
  51. Sekiya K, Nakagawa H (1983) Cytogenetics of Xenopus laevis. I. G-banding pattern of Xenopus laevis chromosomes. Experientia 39:786–787Google Scholar
  52. Sims SH, Macgregor HC, Pellat PS, Horner HA (1984) Chromosome 1 in crested and marbled newts (Triturus). An extraordinary case of heteromorphism and independent chromosome evolution. Chromosoma 89:169–185Google Scholar
  53. Sinclair JH, Brown DD (1971) Retention of common nucleotide sequences in the ribosomal deoxyribonucleic acid of eukaryotes and some of their physical characteristics. Biochemistry 10:2761–2769Google Scholar
  54. Stern R (1972) Satellite DNAs of Xenopus mulleri. Carnegie Inst Wash Yearb 71:22Google Scholar
  55. Stock AD, Mengden GA (1975) Chromosome banding pattern conservatism in birds and nonhomology of chromosome banding patterns between birds, turtles, snakes and amphibians. Chromosoma 50:69–77Google Scholar
  56. Szabo P, Lee MR, Elder FB, Prensky W (1978) Localization of 5S RNA and rRNA genes in the Norway rat. Chromosoma 65:161–172Google Scholar
  57. Thiery JP, Macaya G, Bernardi G (1976) An analysis of eukaryotic genomes by density gradient centrifugation. J Mol Biol 108:219–235Google Scholar
  58. Tymowska J, Fischberg M (1982) A comparison of the karyotype, constitutive heterochromatin, and nucleolar organizer regions of the new tetraploid species Xenopus epitropicalis Fischberg and Picard with those of Xenopus tropicalis Gray (Anura, Pipidae), Cytogenet Cell Genet 34:149–157Google Scholar
  59. Vitelli L, Batistoni R, Andronico F, Nardi I, Barsacchi-Pilone G (1982) Chromosomal localization of 18S + 28S and 5S ribosomal RNA genes in evolutionary diverse anuran amphibians. Chromosoma 84:475–491Google Scholar
  60. Ward DC, Reich E, Goldberg IH (1965) Base specifity in the interaction of polynucleotides with antibiotic drugs. Science 149:1259–1263Google Scholar
  61. Weisblum B (1973) Fluorescent probes of chromosomal DNA structure: three classes of acridines. Cold Spring Harbor Symp Quant Biol 38:441–449Google Scholar
  62. Wolf K, Quimby MC (1964) Amphibian cell culture: permanent cell line from the bullfrog (Rana catesbeiana). Science 114:1578–1580Google Scholar

Copyright information

© Springer-Verlag 1987

Authors and Affiliations

  • M. Schmid
    • 1
  • L. Vitelli
    • 2
  • R. Batistoni
    • 2
  1. 1.Department of Human GeneticsUniversity of WürzburgWürzburgFederal Republic of Germany
  2. 2.Istituto di Istologia e EmbriologiaUniversity of PisaPisaItaly

Personalised recommendations