Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Research Paper
  • Published:

Regeneration of Fertile Plants from Protoplasts of Elite Inbread Maize.

Bio/Technology volume 7, pages 581–587 (1989)Cite this article

Abstract

We have obtained friable, non-mucilaginous embryogenic callus from cultured immature embryos of an elite maize inbred line derived from Iowa stiff stalk synthetic germplasm. This callus was used to generate embryogenic cell suspensions containing compact clumps of densely cytoplasmic cells. Plants were readily regenerated from the suspensions, which were also an excellent source of protoplasts. The suspension cultures maintained their plant regeneration capacity through many months. The cultures were cryopreserved in liquid nitrogen and embryogenic callus and suspension cultures were routinely recovered from cryopreserved cells. Protoplasts were isolated in large numbers from both primary and cryopreserved suspension cultures. Cultured protoplasts divided and formed embryogenic callus both when plated in agarose and on nurse cultures. Hundreds of plants were regenerated from protoplast-derived calli. Plants with viable progeny were obtained although many of the regenerates showed morphological and reproductive abnormalities.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Potrykus, I., Harms, C.T., Lörz, H. and Thomas, E. 1977. Callus formation from stem protoplasts of corn (Zea >mays L.). Molec. Gen. Genet. 156:347–350.

    Article  CAS  Google Scholar 

  2. Potrykus, I., Harms, C.T. and Lörz, H. 1979. Callus formation from cell culture protoplasts of corn (Zea >mays L.). Theor. Appl. Genet. 54:209–214.

    Article  CAS  Google Scholar 

  3. Sheridan, W.F. 1982. Black Mexican Sweet Corn: its use for tissue cultures, p. 385–388. In: Maize for Biological Research. W.F. Sheridan (Ed.). Plant Molecular Biology Association, Charlottesville, Virginia.

    Google Scholar 

  4. Chourey, P.S., and Zurawski, D.B. 1981. Callus formation from protoplasts of a maize cell culture. Theor. Appl. Genet. 59:341–344.

    Article  CAS  Google Scholar 

  5. Fromm, M., Taylor, L.P., and Walbot, V. 1985. Expression of genes transferred into monocot and dicot plant cells by electroporation. Proc. Natl. Acad. Sci. USA 82:5824–5828.

    Article  CAS  Google Scholar 

  6. Vasil, I.K. 1987. Developing cell and tissue culture systems for the improvement of cereal and grass crops. J. Plant Physiol. 128:193–218.

    Article  Google Scholar 

  7. Abdullah, R., Cocking, E.C. and Thompson, J.A. 1986. Efficient plant regeneration from rice protoplasts through somatic embryogenesis. Bio/Technology 4:1087–1090.

    Google Scholar 

  8. Toriyama, K., Hinata, K. and Sasaki, T. 1986. Haploid and diploid plant regeneration from protoplasts of anther callus in rice. Theor. Appl. Genet. 73:16–19.

    Article  CAS  Google Scholar 

  9. Yamada, Y., Yang, Z.Q. and Thang, D.T. 1986. Plant regeneration from protoplast-derived callus of rice (Oryza saliva L.). Plant Cell Rep. 4:85–88.

    Article  Google Scholar 

  10. Kyozuka, J., Hayashi, Y. and Shimamoto, K. 1987. High frequency plant regeneration from rice protoplasts by novel nurse culture methods. Molec. Gen. Genet. 206:408–413.

    Article  CAS  Google Scholar 

  11. Srinivasan, C. and Vasil, I.K. 1986. Plant regeneration from protoplasts of sugarcane (Saccharum officinarum L.). J. Plant Physiol. 126:41–48.

    Article  CAS  Google Scholar 

  12. Rhodes, C.A., Lowe, K.S. and Ruby, K.L. 1988. Plant regeneration from protoplasts isolated from embryogenic maize cell cultures. Bio/Technology 6:56–60.

    Google Scholar 

  13. Vasil, V. and Vasil, I. 1980. Isolation and culture of cereal protoplasts. Part 2: Embryogenesis and plantlet formation from protoplasts of Pennisetum americanum. Theor. Appl. Genet. 56:97–99.

    Article  CAS  Google Scholar 

  14. Lu, C-Y., Vasil, V. and Vasil, I.K. 1981. Isolation and culture of protoplasts of Panicum maximum Jacq. (Guinea grass): Somatic embryogenesis and plantlet formation. Z. Pflanzenphysiol. 104:311–318.

    Article  Google Scholar 

  15. Dalton, S.J. 1988. Plant regeneration from cell suspension protoplasts of Festuca arundinacea Schreb. (tall fescue) and Lolium perenne L. (perennial ryegrass). J. Plant Physiol. 132:170–175.

    Article  Google Scholar 

  16. Horn, M.E., Conger, B.V., and Harms, C.T. 1988. Plant regeneration from protoplasts of embryogenic suspension cultures of orchard-grass (Dactylis gtomerata L.). Plant Cell Rep. 7:371–374.

    Article  CAS  Google Scholar 

  17. Kamo, K.K., Chang, K.L., Lymm, M.E. and Hodges, T.K. 1987. Embryogenic callus formation from maize protoplasts. Planta 172:245–251.

    Article  CAS  Google Scholar 

  18. Vasil, V. and Vasil, I.K. 1987. Plant regeneration from friable embryogenic callus and cell suspension cultures of Zea mays. J. Plant Physiol. 124:399–408.

    Article  Google Scholar 

  19. Imbrie-Milligan, C.W. and Hodges, T.K. 1986. Microcallus formation from maize protoplasts prepared from embryogenic callus. Planta 168:395–401.

    Article  CAS  Google Scholar 

  20. Ludwig, S.R., Somers, D.A., Peterson, W.L., Pohlman, R.F., Zarowits, M.A., Gengenbach, B.G. and Messing, J. 1985. High frequency callus formation from maize protoplasts. Theor. Appl. Genet. 71:344–350.

    Article  CAS  Google Scholar 

  21. Cai, Q., Kuo, C., Qian, Y., Jiong, R. and Zhou, Y. 1988. Somatic embryogenesis and plant regeneration from protoplast of maize (Zea mays L.), p. 120. In: Progress in Plant Protoplast Research; Proc. 7th Int. Protoplast Symp., Wageningen, The Netherlands, 1987. K.J. Puite et al. (Eds). Kluwer, Dordrecht.

    Google Scholar 

  22. Imbrie-Milligan, C.W., Kamo, K.K. and Hodges, T.K. 1987. Micro-callus growth from maize protoplasts. Planta 171:58–64.

    Article  CAS  Google Scholar 

  23. Rhodes, C.A., Pierce, D.A., Mettler, I.J., Mascarenhas, D. and Detmer, J.J. 1988. Genetically transformed maize plants from protoplasts. Science 240:204–207.

    Article  CAS  Google Scholar 

  24. Prioli, L.M. and Söndahl, M.R. 1989. Plant regeneration and recovery of fertile plants from protoplasts of maize (Zea mays L.). Bio/Technology 7:xxx–xxx.

    Google Scholar 

  25. Shillito, R.D. 1978. Problems of auxotrophic mutant isolation. PhD. Thesis, University of Leicester, England.

    Google Scholar 

  26. Ulrich, J.M., Finkle, B.J., Mackey, B.E., Schaeffer, G.W. and Sharpe, F. Jr. 1984. Responses of six rice callus cultures to deep-frozen temperatures. Crop Sci. 24:82–85.

    Article  Google Scholar 

  27. Withers, L.A. and King, P.J. 1979. Proline, a novel cryoprotectant for the freeze-preservation of cultured cells of Zea mays. Plant Physiol. 64:675–678.

    Article  CAS  Google Scholar 

  28. Kuang, V.D., Shamina, Z.B. and Butenko, R.G. 1983. Use of nurse tissue culture to obtain clones from cultured cells and protoplasts of corn. Fiziol. Rast. 30:803–812.

    Google Scholar 

  29. Smith, J.A., Green, C.E. and Gengenbach, B.G. 1984. Feeder layer support of low density populations of Zea mays L. suspension cells. Plant Sci. Lett. 36:67–72.

    Article  Google Scholar 

  30. Carswell, G.K., Johnson, C.M., Shillito, R.D. and Harms, C.T. 1989. Acetyl-salicylic acid promotes colony formation from protoplasts of elite maize inbreds. Plant Cell. Rep. Submitted.

  31. Lee, M. and Phillips, R.L. 1987. Genetic variants in progeny of regenerated maize plants. Genome 29:834–838.

    Article  Google Scholar 

  32. Everett, N.P., Wach, M.J. and Ashworth, D.J. 1985. Biochemical markers of embryogenesis in tissue cultures of the maize inbred B73. Plant Sci. 41:133–140.

    Article  CAS  Google Scholar 

  33. Lowe, K., Barnes-Taylor, D., Ryan, P. and Paterson, K.E. 1985. Plant regeneration via organogenesis and embryogenesis in the maize inbred line B73. Plant Sci. 41:125–132.

    Article  Google Scholar 

  34. Green, C.E. 1982. Somatic embryogenesis and plant regeneration from the friable callus of Zea mays, p. 107–108. In: Plant Tissue Culture 1982. A. Fujiwara (Ed.). Maruzen, Tokyo.

    Google Scholar 

  35. Armstrong, C.L. and Green, C.E. 1984. Establishment and maintenance of friable, embryogenic callus and the involvement of L-proline. Planta 164:207–214.

    Article  Google Scholar 

  36. Vasil, V., Vasil, I.K. and Lu, C. 1984. Somatic embryogenesis in long-term callus cultures of Zea mays L. (Gramineae). Amer. J. Bot. 71:158–161.

    Article  Google Scholar 

  37. Tomes, D.T. and Smith, O.S. 1985. The effect of parental genotype on initiation of embryogenic callus from elite maize (Zea >mays L.) germplasm. Theor. Appl. Genet. 70:505–509.

    Article  CAS  Google Scholar 

  38. Reece, S.L. and Maddox, S.E. 1987. Cryopreservation of embryogenic maize suspension cultures. In Vitro 24:86.

    Google Scholar 

  39. Kao, K.N. and Michayluk, M.R. 1975. Nutrient requirements for growth of Vicia hajastana cells and protoplasts at very low population density in liquid media. Planta 126:105–110.

    Article  CAS  Google Scholar 

  40. Chen, W.H., Davey, M.R., Power, J.B. and Cocking, E.C. 1988. Sugarcane protoplasts: factors affecting division and plant regeneration. Plant Cell Rep. 7:344–347.

    Article  CAS  Google Scholar 

  41. Green, C.E. and Phillips, R.L. 1975. Plant regeneration from tissue cultures of maize. Crop. Sci. 15:417–421.

    Article  Google Scholar 

  42. Duncan, D.R., Williams, M.E., Zehr, B.E. and Widholm, J.M. 1985. The production of callus capable of plant regeneration from immature embryos of numerous Zea >mays genotypes. Planta 165:322–332.

    Article  CAS  Google Scholar 

  43. Harms, C.T., Lörz, H. and Potrykus, I. 1976. Regeneration of plantlets from callus cultures of Zea mays. Z. Pflanzenzüchtg. 77:347–351.

    Google Scholar 

  44. Kamo, K.K. and Hodges, T.K. 1986. Establishment and characterization of long-term embryogenic maize (Zea mays) callus and suspension cultures. Plant Sci. 45:111–118.

    Article  Google Scholar 

  45. Duncan, D.R. and Widholm, J.M. 1988. Improved plant regeneration from maize callus cultures using 6-benzylaminopurine. Plant Cell Rep. 7:452–455.

    CAS  PubMed  Google Scholar 

  46. Edallo, S., Zucchinalli, C., Perenzin, M. and Salamini, F. 1981. Chromosomal variation and frequency of spontaneous mutation associated with in vitro culture and plant regeneration in maize. Maydica 26:39–56.

    Google Scholar 

  47. Chu, C.C., Wang, C.C., Sun, C.S., Hsu, C., Yin, K.G., Chu, C.Y. and Bi, F.Y. 1975. Establishment of an efficient medium for anther culture of rice through comparative experiments on the nitrogen sources. Scientia Sinica 18:659–668.

    Google Scholar 

  48. Murashige, T. and Skoog, F. 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant 15:473–497.

    Article  CAS  Google Scholar 

  49. Gamborg, O.L., Miller, R.A. and Ojima, L. 1968. Nutrient requirements of suspension cultures of soybean root cells. Exp. Cell Res. 50:151–158.

    Article  CAS  Google Scholar 

  50. Harms, C.T. and Potrykus, I. 1978. Fractionation of plant protoplast types by iso-osmotic density gradient centrifugation. Theor. Appl. Genet. 53:57–63.

    Article  CAS  Google Scholar 

  51. Shillito, R.D., Paszkowski, J. and Potrykus, I. 1983 Agarose plating and a bead type culture technique enable and stimulate development of protoplast-derived colonies in a number of plant species. Plant Cell Rep. 2:244–247.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Ciba-Geigy Corporation, Agricultural Biotechnology Research Unit, P.O. Box 12257, Research Triangle Park, North Carolina, 27709

    Raymond D. Shillito, Gleta K. Carswell, Catherine M. Johnson, Joseph J. DiMaio & Christian T. Harms

Authors
  1. Raymond D. Shillito
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Gleta K. Carswell
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Catherine M. Johnson
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Joseph J. DiMaio
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Christian T. Harms
    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

Shillito, R., Carswell, G., Johnson, C. et al. Regeneration of Fertile Plants from Protoplasts of Elite Inbread Maize.. Nat Biotechnol 7, 581–587 (1989). https://doi.org/10.1038/nbt0689-581

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt0689-581

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing