, Volume 91, Issue 1, pp 28–38 | Cite as

Chromosomal and nuclear distribution of the HindIII 1.9-kb human DNA repeat segment

  • Laura Manuelidis
  • David C. Ward


A human interspersed repetitive DNA cloned in pBR322, the HindIII 1.9-kb (kilobase pair) sequence, was labeled with biotinylated dUTP and hybridized to acid-fixed chromosomes and paraformaldehyde-fixed whole cells in situ. Using our most sensitive detection techniques this probe highlighted on the order of 200 discrete loci, in punctate or banded arrays, that resembled a Giemsa-dark band pattern on chromosome arms. Interphase cells also displayed many discrete punctate spots of hybridization along chromosome fibers. The ubiquitous Alu sequence repeat also appeared to be concentrated in specific regions of the chromosome and predominantly highlighted Giemsa-light bands. Centromeric or ribosomal spacer DNA repeats used as controls in all studies gave the expected hybridization profiles and showed no non-specific labeling of chromosome arms. Cohesive groups of centromeric DNA arrays and rDNA clusters were observed in interphase nuclei. Refinements in methods for detecting biotin-labeled probes in situ were developed during these studies and calculations indicated that about 20 kb or more of the 1.9-kb repeat were present at each hybridization site. The chromosomal distribution of the 1.9-kb repeat suggests that this sequence may reflect, or participate in defining, ordered structureal domains along the chromosome.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Adams J, Kaufman RE, Dretschner PJ, Harrison M, Nienhuis AW (1980) A family of long reiterated DNA sequences, one copy of which is next to the human beta globin gene. Nucleic Acids Res 8:6113–6129Google Scholar
  2. Alberstson DG (1984) Localization of the ribosomal genes in Caenorhabditis elegans chromosomes by in situ hybridization using biotin-labeled probes. EMBO J 3:1227–1234Google Scholar
  3. Bogenhagen DF, Sakonja S, Brown DD (1980) A control region in the center of the 5S RNA gene directs specific initiation of transcription: II. The 3′ border of the region. Cell 19:27–35Google Scholar
  4. Brigati DJ, Myerson D, Leary JJ, Spalholz B, Travis SZ, Fong CKY, Hsiung GD, Ward DC (1983) Detection of viral genomes in cultured cells and paraffin embedded tissue sections using biotin-labeled hybridization probes. Virology 126:32–50Google Scholar
  5. Capco DC, Krochmalnic G, Penman A (1984) A new method for preparing embeddment-free sections for transmission electron microscopy: applications to the cytoskeletal framework and other three-dimensional networks. J Cell Biol 98:1878–1885Google Scholar
  6. Denhardt DT (1966) A membrane filter technique for the detection of complementary DNA. Biochim Biophys Res Comm 23:183–184Google Scholar
  7. DiGiovanni L, Haynes SR, Misra R, Jelinek WR (1983) Kpn I family of long-dispersed repeated DNA sequences of man: Evidence for entry into genomic DNA of DNA copies of poly(A)-terminated Kpn I RNAs. Proc Natl Acad Sci USA 80:6533–6537Google Scholar
  8. Doolittle WF, Sapienza C (1980) Selfish genes, the phenotype paradigm and genome evolution. Nature 284:601–603Google Scholar
  9. Evans H, Buckland R, Pardue ML (1966) Location of the genes coding for 18S and 28S ribosomal RNA in the human genome. Chromosoma 48:405–426Google Scholar
  10. Gallyas F, Gores T, Merchenthaler I (1982) High-grade intensification of the end product of the diaminobenzidine reaction for peroxidase histochemistry. J Histochem Cytochem 30:183–184Google Scholar
  11. Georgiev GP, Kramerov DA, Ryskov AP, Skryabin KG, Lukanidin EM (1982) Dispersed repetitive sequences in eukaryotic genomes and their possible biological significance. Cold Spring Harbor Symp Quant Biol 47:1109–1121Google Scholar
  12. Gerhard DS, Kawasaki ES, Bancroft FC, Szabo P (1981) Localization of a unique gene by direct hybridization in situ. Proc Natl Acad Sci USA 78:3755–3759Google Scholar
  13. Goldman MA, Holmquist GP, Gray MC, Caston LA, Nag A (1984) Replication timing of genes and middle repetitive sequences. Science 224:686–692Google Scholar
  14. Harper ME, Saunders GF (1981) Localization of single copy DNA sequences on G-banded human chromosomes by in situ hybridization. Chromosoma 83:431–439Google Scholar
  15. Holmquist G, Gray M, Porter T, Jordan J (1982) Characterization of Giemsa-dark and light-band DNA. Cell 31:121–129Google Scholar
  16. Inoué S (1981) Video image processing greatly enhances contrast, quality and speed in polarization-based microscopy. J Cell Biol 89:346–356Google Scholar
  17. Jelinek WR, Toomey TP, Leinwand L, Duncan CH, Biro PA, Choudary PV, Weissman SM, Rubin CM, Houck CM, Deininger PL, Schmid CW (1980) Ubiquitous interspersed repeated sequences in mammalian genomes. Proc Natl Acad Sci USA 77:1398–1402Google Scholar
  18. Kole LB, Haynes SR, Jelinek WR (1983) Discrete and heterogeneous high molecular weight RNAs complementary to a long dispersed repeat family (a possible transposon) of human DNA. J Mol Biol 165:257–286Google Scholar
  19. Langer PR, Waldrop AA, Ward DC (1981) Enzymatic synthesis of biotin-labeled polynucleotides: novel nucleic acid affinity probes. Proc Natl Acad Sci USA 78:6633–6637Google Scholar
  20. Leary JJ, Brigati DJ, Ward DC (1983) Rapid sensitive colorimetric method for visualizing biotin-labeled DNA probes hybridized to DNA or RNA immobilized on nitrocellulose: Bio-blots. Proc Natl Acad Sci USA 80:4045–4049Google Scholar
  21. Lerman MI, Thayer RE, Singer MF (1983) Kpn I family of long interspersed repeated DNA sequences in primates: polymorphism of family members and evidence for transcription. Proc Natl Acad Sci USA 80:3966–3970Google Scholar
  22. Maio JJ, Brown FL, McKenna WG, Musich PR (1981) Toward a molecular paleontology of primate genomes. II. The Kpn I families of alphoid DNAs. Chromosoma 83:127–144Google Scholar
  23. Manuelidis L, (1980) Novel classes of mouse repeated DNAs. Nucleic Acids Res 8:3247–3258Google Scholar
  24. Manuelidis L (1982a) Repeated DNA sequences and nuclear structure. In: Dover GA, Flavell RB (eds) Genome evolution. Academic Press, New York, pp 261–285Google Scholar
  25. Manuelidis L (1982b) Nucleotide sequence definition of a major human repeated DNA, the HindIII 1.9-kb family. Nucleic Acids Res 10:3211–3219Google Scholar
  26. Manuelidis L (1984a) Different CNS cell types display distinct and non-random arrangements of satellite DNA sequences. Proc Natl Acad Sci USA 81:3123–3127Google Scholar
  27. Manuelidis L (1984b) Active nucleolus organizers are precisely positioned in adult central nervous system cells but not in neuroectodermal tumor cells. J Neuropathol Exp Neurol 43:225–241Google Scholar
  28. Manuelidis L, Biro PA (1982) Genomic representation of the Hind-III 1.9-kb repeated DNA. Nucleic Acids Res 10:3221–3239Google Scholar
  29. Manuelidis L, Manuelidis EE (1979) Surface growth characteristics of defined normal and neoplastic neuroectodermal cells in vitro. In: Zimmerman HM, (ed) Progress in neuropathology, vol. 4, Raven Press, New York, pp 235–285Google Scholar
  30. Manuelidis L, Langer-Safer PR, Ward DC (1982) High resolution mapping of satellite DNA using biotin-labeled DNA probes. J Cell Biol 95:619–625Google Scholar
  31. Marsden MPF, Laemmli UK (1979) Metaphase chromosome structure: evidence for a radial loop model. Cell 17:849–858Google Scholar
  32. Martin SL, Voliva CF, Burton FH, Edgell MH, Hutchison CA III (1984) A large interspersed repeat found in mouse DNA contains a long open reading frame that evolves as if it encodes a protein. Proc Natl Acad Sci USA 81:2308–2312Google Scholar
  33. Newman GR, Jasani B, Williams ED (1983a) The visualization of trace amounts of diaminobenzidine (DAB) polymer by a novel gold-sulphide-silver method. J Microsc 132:1–2Google Scholar
  34. Newman GR, Jasani B, Williams ED (1983b) Metal compound intensification of the electron-density of diaminobenzidine. J Histochem Cytochem 31:1430–1434Google Scholar
  35. Pan J, Elder JT, Duncan CH, Weissman SM (1981) Structural analysis of interspersed repetitive polymerase transcription units in human DNA. Nucleic Acids Res 9:1151–1170Google Scholar
  36. Potter SS (1984) Rearranged sequences of a human Kpn I element. Proc Natl Acad Sci USA 81:1012–1016Google Scholar
  37. Rubin CM, Houck CM, Deininger PL, Friedmann T, Schmid CW (1980) Partial nucleotide sequence of the 300 nucleotide interspersed repeated human DNA sequences. Nature 284:372–374Google Scholar
  38. Sakonju S, Bogenhagen DF, Brown DD (1980) A control region in the center of the 5S RNA gene directs specific initiation of transcription. I. The 5′ border of the region. Cell 19:13–25Google Scholar
  39. Schmeckpeper BJ, Willard HF, Smith KD (1981) Isolation and characterization of cloned human DNA fragments carrying reiterated sequences common to both autosomes and the X-chromosome. Nucleic Acids Res 9:1853–1872Google Scholar
  40. Sedat J, Manuelidis L (1978) A direct approach to the structure of eukaryotic chromosomes. Cold Spring Harbor Symp Quant Biol 42:331–350Google Scholar
  41. Shafit-Zagardo B, Maio JJ, Brown FL (1982) Kpn I families of long interspersed repetitive DNAs in human and other primate genomes. Nucleic Acids Res 10:3175–3179Google Scholar
  42. Shafit-Zagardo B, Brown FL, Zavodny PJ, Maio JJ (1983) Transcription of the Kpn I families of long interspersed DNAs in human cells. Nature 304:277–280Google Scholar
  43. Sharp PA (1983) Conversion of RNA to DNA in mammals: Alulike elements and pseudogenes. Nature 301:471–472Google Scholar
  44. Singer MF (1982) Highly repeated sequences in mammalian genomes. Int Rev Cytol 76:67–112Google Scholar
  45. Singer MF, Thayer RE, Grimaldi G, Fanning TG (1983) Homology between the Kpn I primate and Bam HI (MIF-1) rodent families of long interspersed repeated sequences. Nucleic Acids Res 11:5739–5745Google Scholar
  46. Thayer RE, Singer ML (1983) Interruption of an α-satellite array by a short member of the Kpn I family of interspersed, highly repeated monkey DNA sequences. Mol Cell Biol 3:967–973Google Scholar
  47. Varley KM, MacGregor HC, Euba HP (1980) Satellite DNA is transcribed on lampbrush chromosomes. Nature 283:686–688Google Scholar
  48. Willard HF, Smith KD, Sutherland J (1983) Isolation and characterization of a major tandem repeat family from the human X chromosome. Nucleic Acids Res 11:2017–2033Google Scholar
  49. Wu JC, Manuelidis L (1980) Sequence definition and organization of a human repeated DNA. J Mol Biol 142:363–386Google Scholar
  50. Yang TP, Hansen SK, Oishi KK, Ryder DA, Hamkalo BA (1982) Characterization of a cloned repetitive DNA sequence concentrated on the human X chromosome. Proc Natl Acad Sci USA 79:6593–6597Google Scholar

Copyright information

© Springer-Verlag 1984

Authors and Affiliations

  • Laura Manuelidis
    • 1
  • David C. Ward
    • 2
  1. 1.Section of NeuropathologyYale University School of MedicineNew HavenUSA
  2. 2.Department of Human Genetics and Molecular Biophysics-BiochemistryYale University School of MedicineNew HavenUSA

Personalised recommendations