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Planta

, Volume 224, Issue 1, pp 72–82 | Cite as

Changes in chloroplast DNA during development in tobacco, Medicago truncatula, pea, and maize

  • Jeffrey M. Shaver
  • Delene J. Oldenburg
  • Arnold J. BendichEmail author
Original Article

Abstract

We examined the DNA from chloroplasts obtained from young and fully expanded leaves of tobacco (Nicotiana tabacum L.), Medicago truncatula, pea (Pisum sativum L.), and maize (Zea mays L.). The changes in plastid DNA content and structure were monitored by four independent methods: 4′,6-diamidino-2-phenylindole (DAPI) staining with intact chloroplasts, in situ DAPI staining of cytological sections, ethidium bromide staining at the single-molecule level after exhaustive deproteinization of lysed chloroplasts, and pulsed-field gel electrophoresis. During leaf development, we found a decline of chloroplast DNA (cpDNA) in all four plants. For tobacco, for which plants can readily be regenerated from somatic cells, cpDNA persisted longer than in the other three plants. We also found a striking progression from complex multigenomic DNA molecules to simple subgenomic molecules during plastid development. Although the decrease in molecular size and complexity paralleled the decrease in DNA content per plastid, 6% of the chloroplasts in a fully expanded tobacco leaf still contained DNA in complex branched structure, whereas no such complex structures were found in mature leaves for the hard-to-regenerate maize.

Keywords

Chloroplast DNA DAPI Development Fluorescence microscopy Pulsed-field gel electrophoresis Regeneration 

Abbreviations

cpDNA

Chloroplast DNA

DAPI

4′,6-diamidino-2-phenylindole

IR

Inverted repeat

Notes

Acknowledgements

The project was supported by the National Research Initiative of the USDA Cooperative State Research, Education and Extension Service, grant number#2002-35301-12021. We thank Marc Schleh for conducting the maize assays and Doug Ewing for assistance with growing plants.

References

  1. Backert S, Dörfel P, Börner T (1995) Investigation of plant organellar DNAs by pulsed-field gel electrophoresis. Curr Genet 28:390–399PubMedCrossRefGoogle Scholar
  2. Baumgartner BJ, Rapp JC, Mullet JE (1989) Plastid transcription activity and DNA copy number increase early in barley chloroplast development. Plant Physiol 89:1011–1018PubMedGoogle Scholar
  3. Bendich A (1987) Why do chloroplasts and mitochondria contain so many copies of their genome? Bioessays 6:279–282PubMedCrossRefGoogle Scholar
  4. Bendich A (1991) Moving pictures of DNA released upon lysis from bacteria, chloroplasts, and mitochondria. Protoplasma 160:121–130CrossRefGoogle Scholar
  5. Bendich A (2004) Circular chloroplast chromosomes: the grand illusion. Plant Cell 16:1661–1666PubMedCrossRefGoogle Scholar
  6. Bookjans G, Stummann BM, Henningsen KW (1984) Preparation of chloroplast DNA from pea plastids isolated in a medium of high ionic strength. Anal Biochem 141:244–247PubMedCrossRefGoogle Scholar
  7. Deng X-W, Wing RA, Gruissem W (1989) The chloroplast genome exists in multimeric forms. Proc Natl Acad Sci USA 86:4156–4160PubMedCrossRefGoogle Scholar
  8. Fujie M, Kuroiwa H, Kawano S, Mutoh S, Kuroiwa T (1994) Behavior of organelles and their nucleoids in the shoot apical meristem during leaf development in Arabidopsis thaliana L. Planta 194:395–405CrossRefGoogle Scholar
  9. Kuroiwa T (1991) The replication, differentiation, and inheritance of plastids with emphasis on the concept of organelle nuclei. Int Rev Cytol 128:1–62Google Scholar
  10. Kuroiwa T, Suzuki T (1980) An improved method for the demonstration of the in situ chloroplast nuclei in higher plants. Cell Struct Funct 5:195–197Google Scholar
  11. Kuroiwa T, Suzuki T, Ogawa K, Kawano S (1981) The chloroplast nucleus: distribution, number, size, and shape, and a model for multiplication of the chloroplast genome during chloroplast development. Plant Cell Physiol 22:381–396Google Scholar
  12. Lamppa G, Bendich A (1979) Changes in chloroplast DNA levels during development of pea (Pisum sativum). Plant Physiol 64:126–130PubMedGoogle Scholar
  13. Lamppa GK, Elliot LV, Bendich AJ (1980) Changes in chloroplast number during pea leaf development: an analysis of a protoplast population. Planta 148:437–443CrossRefGoogle Scholar
  14. Li B, Huang W, Bass T (2003) Shoot production per responsive leaf explant increases exponentially with explant organogenic potential in Nicotiana species. Plant Cell Rep 22:231–238CrossRefPubMedGoogle Scholar
  15. Luo Y, Koop H-U (1997) Somatic embryogenesis in cultured immature zygotic embryos and leaf protoplasts of Arabidopsis thaliana ecotypes. Planta 202:387–396CrossRefPubMedGoogle Scholar
  16. Miyamura S, Nagata T, Kuroiwa T (1986) Quantitative fluorescence microscopy on dynamic changes of plastid nucleoids during wheat development. Protoplasma 133:66–72CrossRefGoogle Scholar
  17. Nishimura Y, Misumi O, Kato K, Inada N, Higashiyama T, Momoyama Y, Kuroiwa T (2002) An mt+ gamete-specific nuclease that targets mtchloroplasts during sexual reproduction in C. reinhardtii. Genes Dev 16:1116–1128PubMedCrossRefGoogle Scholar
  18. Ogihara Y, Isono K, Kojima K, Endo A, Hanaoka M, Shiina T, Terachi T, Utsugi S, Murata M, Mori N, Takumi S, Ikeo K, Gojobori T, Murai R, Murai K, Matsuoka Y, Ohnishi Y, Tajiri H, Tsunewaki K (2002) Structural features of a wheat plastome as revealed by complete sequencing of chloroplasts. Mol Genet Genome 266:740–746CrossRefGoogle Scholar
  19. Oldenburg D, Bendich A (1996) Size and structure of replicating mitochondrial DNA in cultured tobacco cells. Plant Cell 8:447–461CrossRefPubMedGoogle Scholar
  20. Oldenburg D, Bendich A (1998) The structure of mitochondrial DNA from the liverwort, Marchantia polymorpha. J Mol Biol 276:745–758CrossRefPubMedGoogle Scholar
  21. Oldenburg D, Bendich A (2004a) Most chloroplast DNA of maize seedlings in linear molecules with defined ends and branched forms. J Mol Biol 335:953–970CrossRefPubMedGoogle Scholar
  22. Oldenburg D, Bendich A (2004b) Changes in the structure of DNA molecules and the amount of DNA per plastid during chloroplast development in maize. J Mol Biol 344:1311–1330CrossRefPubMedGoogle Scholar
  23. Rowan B, Oldenburg D, Bendich A (2004) The demise of chloroplast DNA in Arabidopsis. Curr Genet 46:176–181PubMedCrossRefGoogle Scholar
  24. Scott NS, Possingham JV (1980) Chloroplast DNA in expanding spinach leaves. J Exper Bot 31:1081–1092CrossRefGoogle Scholar
  25. Seyer P, Kowallik KV, Herrman RG (1981) A physical map of Nictotiana tabacum plastid DNA including the location of structural genes for ribosomal RNAs and the large subunit of ribulose bisphosphate carboxylase/oxygenase. Curr Genet 3:189–204CrossRefGoogle Scholar
  26. Sodmergen, Kawano S, Tano S, Kuroiwa T (1991) Degradation of chloroplast DNA in second leaves of rice (Oryza sativa) before leaf yellowing. Protoplasma 160:89–98CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Jeffrey M. Shaver
    • 1
  • Delene J. Oldenburg
    • 1
  • Arnold J. Bendich
    • 1
    Email author
  1. 1.Department of BiologyUniversity of WashingtonSeattleUSA

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