Skip to main content
SpringerLink
Log in

Structural analysis of two length variants of the rDNA intergenic spacer from Eruca sativa

  • Research Article
  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

Restriction enzyme analysis of the rRNA genes of Eruca sativa indicated the presence of many length variants within a single plant and also between different cultivars which is unusual for most crucifers studied so far. Two length variants of the rDNA intergenic spacer (IGS) from a single individual E. sativa (cv. Itsa) plant were cloned and characterized. The complete nucleotide sequences of both the variants (3 kb and 4 kb) were determined. The intergenic spacer contains three families of tandemly repeated DNA sequences denoted as A, B and C. However, the long (4 kb) variant shows the presence of an additional repeat, denoted as D, which is a duplication of a 224 bp sequence just upstream of the putative transcription initiation site. Repeat units belonging to the three different families (A, B and C) were in the size range of 22 to 30 bp. Such short repeat elements are present in the IGS of most of the crucifers analysed so far. Sequence analysis of the variants (3 kb and 4 kb) revealed that the length heterogeneity of the spacer is located at three different regions and is due to the varying copy numbers of repeat units belonging to families A and B. Length variation of the spacer is also due to the presence of a large duplication (D repeats) in the 4 kb variant which is absent in the 3 kb variant. The putative transcription initiation site was identified by comparisons with the rDNA sequences from other plant species.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Agnihotri A, Gupta V, Lakshmikumaran M, Shivanna KR, Prakash S, Jagannathan V: Production of Eruca-Brassica hybrids by embryo rescue. Plant Breed 104: 281–289 (1990).

    Google Scholar 

  2. Appels R, Dvorak J: The wheat ribosomal DNA spacer region; its structure and variation in populations and among species. Theor Appl Genet 63: 337–348 (1982).

    Google Scholar 

  3. Barker RF, Harberd NP, Jarvis MG, Flavell RB: Structure and evolution of the intergenic region of a ribosomal DNA repeat unit of wheat. J Mol Biol 201: 1–17 (1988).

    Google Scholar 

  4. Bennet RI, Smith AG: The complete nucleotide sequence of the intergenic spacer of an rDNA operon from Brassica oleracea and its comparison with other crucifers. Plant Mol Biol 16: 1095–1098 (1991).

    Google Scholar 

  5. Coen ES, Dover GA: Multiple Pol I initiation sequences of rDNA spacers of Drosophila melanogaster. Nucl Acids Res 10: 7017–7026 (1982).

    Google Scholar 

  6. Cordesse F, Second G, Delseny M: Ribosomal gene spacer length variability in cultivated and wild rice species. Theor Appl Genet 79: 81–88 (1990).

    Google Scholar 

  7. Cordesse F, Grellet F, Reddy AS, Delseny M: Genome specificity of rDNA spacer fragments from Oryza sativa L. Theor Appl Genet 83: 864–870 (1992).

    Google Scholar 

  8. Cordesse F, Cooke R, Tremousaygue D, Grellet F, Delseny M: Fine structure and evolution of the rDNA intergenic spacer in rice and other cereals. J Mol Evol 36: 369–379 (1993).

    Google Scholar 

  9. Dellaporta SL, Wood J, Hicks JB. Maize DNA miniprep. In: Molecular Biology of Plants: A Laboratory Course Manual, pp. 36–37. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1984).

    Google Scholar 

  10. Delcasso-Tremousaygue D, Grellet F, Panabieres F, Ananiev ED, Delseny M: Structural and transcriptional characterization of the external spacer of a ribosomal RNA nuclear gene from a higher plant. Eur J Biochem 172: 767–776 (1988).

    Google Scholar 

  11. Delseny M, Cooke R, Penon P: Sequence heterogeneity in radis nuclear ribosomal RNA genes. Plant Sci Lett 30: 107–119 (1983).

    Google Scholar 

  12. Delseny M, Laroche M, Penon P: Methylation pattern of radish (Raphanus sativus) nuclear ribosomal RNA genes. Plant Physiol 76: 627–632 (1984).

    Google Scholar 

  13. Delseny M, McGrath JM, This P, Chevre AM, Quiros CF: Ribosomal RNA genes in diploid and amphidiploid Brassica and related species: organization, polymorphism, and evolution. Genome 33: 733–744 (1990).

    Google Scholar 

  14. Dover G: Molecular drive: a cohesive mode of species evolution. Nature 299: 111–117 (1982).

    Google Scholar 

  15. Dover G: Molecular drive in multigene families: how biological novelties arise, spread and are assimilated. Trends Genet 2: 159–165 (1986).

    Google Scholar 

  16. Echeverria M, Delcasso-Tremousaygue D, Delseny M: A nuclear protein fraction binding to dA/dT-rich sequences upstream from the radish DNA promoter. Plant J 2: 211–219 (1992).

    Google Scholar 

  17. Ellis THN, Davies DR, Castleton JA, Bedford ID: The organization and genetics of rDNA length variants in peas. Chromosoma 91: 74–81 (1984).

    Google Scholar 

  18. Feinberg AP, Vogelstein B: A technique for radio labelling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 132: 6–13 (1983).

    Google Scholar 

  19. Flavell RB: The structure and control of expression of ribosomal RNA genes. Oxford Surv Plant Mol Cell Biol 3: 252–274 (1986).

    Google Scholar 

  20. Gerlach WL, Bedbrook JR: Cloning and characterization of ribosomal RNA genes from wheat and barley. Nucl Acids Res 7: 1869–1885 (1979).

    Google Scholar 

  21. Grellet F, Delcasso-Tremousaygue D, Delseny M: Isolation and characterization of an unusual repeated sequence from the ribosomal intergenic spacer of the crucifer Sisymbrium irio. Plant Mol Biol 12: 695–706 (1989).

    Google Scholar 

  22. Grimaldi G, Nocera P: Transient expression of Drosophila melanogaster rDNA promoter into cultured Drosophila cells. Nucl Acids Res 14: 6417–6432 (1986).

    Google Scholar 

  23. Gruendler P, Unfried I, Pascher K, Schweizer D: The rDNA intergenic region from Arabidopsis thaliana: structural analysis, intraspecific variation and functional implications. J Mol Biol 221: 1209–1222 (1991).

    Google Scholar 

  24. Grunstein M, Hogness DS: Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene. Proc Natl Acad Sci USA 72: 3961 (1975).

    Google Scholar 

  25. Haltiner-Jones M, Learned RM, Tjian R: Analysis of clustered point mutations in the human ribosomal RNA gene promoter by transient expression in vivo. Proc Natl Acad Sci USA 85: 669–673 (1988).

    Google Scholar 

  26. Katayama S, Hirano HY, Tsukaya H, Naito S, Komeda Y: Analysis of intergenic spacer regions in the nuclear rDNA of Pharbitis nil. Genome 35: 92–97 (1992).

    Google Scholar 

  27. Kavanagh TA, Timmis JN: Heterogeneity in cucumber ribosomal DNA. Theor Appl Genet 72: 337–345 (1986).

    Google Scholar 

  28. Kelly RJ, Johnson RD, Siegel A: Heterogeneity and organization of the ribosomal RNA genes of Cucurbita maxima. Plant Mol Biol 14: 927–933 (1990).

    Google Scholar 

  29. Kohorn BD, Rae PMM: A component of Drosophila RNA polymerase I lies within the rRNA transcription. Nature 304: 179–181 (1983).

    Google Scholar 

  30. Kraft R, Tardiff J, Kruter KS, Leinwand LA: Using miniprep plasmid DNA for sequencing double-stranded templates with Sequenase. Biotechniques 6: 544–546 (1988).

    Google Scholar 

  31. Labhardt P, Reeder RH: Enhancer-like properties of the 60/81 bp elements in the ribosomal gene spacer of Xenopus laevis. Cell 37: 285–289 (1984).

    Google Scholar 

  32. Lassner M, Dvorak J: Preferential homogenisation between adjacent and alternate sub-repeats in wheat rDNA. Nucl Acids Res 14: 5499–5512 (1986).

    Google Scholar 

  33. Luschnig C, Bachmair A, Schweizer D: Intraspecific length heterogeneity of the rDNA-IGR in Arabidopsis thaliana due to homologous recombination. Plant Mol Biol 22: 543–545 (1993).

    Google Scholar 

  34. Malmberg R, Messing J, Sussex I: Molecular Biology of Plants: A Laboratory Course Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1985).

    Google Scholar 

  35. McMullen MD, Hunter B, Phillips RL, Rubenstein I: The structure of the maize ribosomal DNA spacer region. Nucl Acids Res 14: 4953–4969 (1986).

    Google Scholar 

  36. Piller KJ, Baerson S, Polans NO, Kaufman LS: Structural analysis of the short length ribosomal DNA variant from Pisum sativum L. ev. Alaska. Nucl Acids Res 18: 3135–3145 (1990).

    Google Scholar 

  37. Pikaard CS, Reeder RH: Sequence elements essential for function of the Xenopus laevis ribosomal DNA enhancers. Mol Cell Biol 8: 4282–4288 (1988).

    Google Scholar 

  38. Pruitt RE, Meyerowitz EM: Characterization of the genome of Arabidopsis thaliana. J Mol Biol 187: 169–183 (1986).

    Google Scholar 

  39. Rathgeber J, Capesius I: Nucleotide sequence of the intergenic spacer and the 18S ribosomal RNA gene from mustard (Sinapis alba). Nucl Acids Res 18: 1288 (1990).

    Google Scholar 

  40. Sanger F, Nicklen S, Coulson AR: DNA sequencing with chain terminating inhibitors. Proc Natl Acad Sci USA 74: 5463–5467 (1977).

    Google Scholar 

  41. Sollner-Webb B, Wilkinson JA, Roan J, Reeder RH: Nested control regions promote Xenopus ribosomal RNA synthesis by RNA polymerase I. Cell 35: 199–206 (1983).

    Google Scholar 

  42. Sollner-Webb B, Tower J: Transcription of cloned eukaryotic ribosomal RNA genes. Annu Rev Biochem 55: 801–830 (1986).

    Google Scholar 

  43. Taira T, Kato A, Tanifuji S: Difference between two major size classes of carrot rDNA repeating units is due to reiteration of sequences of about 460 bp in the large spacer. Mol Gen Genet 213: 170–174 (1988).

    Google Scholar 

  44. Thompson WF, Flavell RB, Watson JC, Kaufman LS: Chromatin structure and expression of plant ribosomal genes. In: Kahl G (ed). Architecture of Eucaryotic Genes, pp. 385–396. VHC, Weinheim (1988).

    Google Scholar 

  45. Toloczyki C, Feix G: Occurrence of nine homologous repeat units in the external spacer of a nuclear maize rRNA gene unit. Nucl Acids Res 14: 5969–4986 (1986).

    Google Scholar 

  46. Tremousaygue D, Grellet, Delseny M, Delourme R, Renard M: The large spacer of a nuclear ribosomal RNA gene from radish: organization and use as a probe in rapeseed breeding. Theor Appl Genet 75: 289–304 (1988).

    Google Scholar 

  47. Yakura K, Kato A, Tanifuji S: Length heterogeneity in the large spacer of Vicia faba rDNA is due to the differing number of a 325 bp repetitive sequence element. Mol Gen Genet 193: 400–405 (1984).

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Biotechnology, Tata Energy Research Institute, 90 Jor Bagh, 110 003, New Delhi, India

    Malathi Lakshmikumaran & Madan Singh Negi

Authors
  1. Malathi Lakshmikumaran
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Madan Singh Negi
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Lakshmikumaran, M., Negi, M.S. Structural analysis of two length variants of the rDNA intergenic spacer from Eruca sativa . Plant Mol Biol 24, 915–927 (1994). https://doi.org/10.1007/BF00014445

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00014445

Key words

Search

Navigation