Skip to main content
SpringerLink
Log in

Protoplast isolation and regeneration in the marine red alga Porphyra nereocystis

  • Published:
Planta Aims and scope Submit manuscript

Abstract

We have developed a method for isolating viable protoplasts from the blade phase of the epiphytic marine red alga Porphyra nereocystis Anderson, using a two-step enzymatic digestion with commercially available enzymes. The first step uses papain, the second step uses abalone acetone powder. The method is rapid and gives a high yield of viable protoplasts. In liquid culture in enriched seawater medium, the protoplasts can undergo regeneration along three pathways: they directly form filaments resembling the conchocelis phase of Porphyra; they form calli with relatively thick-walled, pigmented cells; and they indirectly form blades from the edges of these calli. Porphyra nereocystis protoplasts also may serve as an alternative propagation method in aquaculture and be useful for studies of cell-wall formation, cell division, and thallus differentiation. They may also be used in somatic selection, somatic hybridization and gene-transfection experiments.

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.

Similar content being viewed by others

Abbreviations

AAP:

abalone acetone powder

PAP:

papain

FDA:

fluorescein diacetate

References

  • Berliner, M.D. (1981) Protoplasts of eukaryotic algae. Int. Rev. Cytol. 73, 1–19

    Google Scholar 

  • Berliner, M.D. (1983) Protoplasts of nonvascular plants. Int. Rev. Cytol. Suppl. 16, 21–31

    Google Scholar 

  • Buggeln, R.G. (1981) Morphogenesis and growth regulators. In: Biology of seaweeds, pp. 627–660, Lobban, C.S., Wynne, M.J., eds. Blackwell, Oxford, UK

    Google Scholar 

  • Burzycki, G.M., Waaland, J.R. (1987) On the position of meiosis in the life history of Porphyra torta (Rhodophyta). Bot. Mar. 30, 5–10

    Google Scholar 

  • Chen, L. C.-M. (1986) Cell development of Porphyra miniata (Rhodophyceae) under axenic culture. Bot. Mar. 29, 435–439

    Google Scholar 

  • Chen, L. C.-M. (1987) Protoplast morphogenesis of Porphyra leucosticta in culture. Bot. Mar. 30, 399–403

    Google Scholar 

  • Cheney, D.P. (1986) Genetic engineering in seaweeds: applications and current status. In: Algal biomass technologies: an interdisciplinary perspective, pp. 22–29, Barclay, W.R., McIntosh, R.P., eds. Beih. Nova Hedwigia No. 83. Cramer, Berlin, Germany

    Google Scholar 

  • Cheney, D.P., Mar, E., Saga, N., van der Meer, J. (1986) Protoplast isolation and cell division in the agar-producing seaweed Gracilaria. J. Phycol. 22, 238–243

    Google Scholar 

  • Dickson, L.G., Waaland, J.R. (1985) Porphyra nereocystis: a dualdaylength seaweed. Planta 165, 548–553

    Google Scholar 

  • Ducreux, G., Kloareg, B. (1988) Plant regeneration from protoplasts of Sphacelaria (Phaeophyceae). Planta 174, 25–29

    Google Scholar 

  • Evans, D.A., Bravo, J.E. (1983) Plant protoplast isolation and culture. Int. Rev. Cytol. Suppl. 16, 33–53

    Google Scholar 

  • Frei, E., Preston, R.D. (1964) Non-cellulosic structural polysaccharides in algal cell walls. II. Association of xylan and mannan in Porphyra umbilicalis Proc. R. Soc. London Ser. B 160, 314–327

    Google Scholar 

  • Fujita, Y., Migita, S. (1985) Isolation and culture of protoplasts from some seaweeds. Bull. Fac. Fish. Nagasaki Univ. 57, 39–45

    Google Scholar 

  • Gretz, M.R., Waaland, J.R. (1989) Biochemistry of cell walls of field collected and cultured Porphyra nereocystis. XIIIth Int. Seaweed Symp., Abstr., p. A10. Vancouver, B.C., Canada

  • Gretz, M.R., Aronson, J.M., Sommerfeld, M.R. (1980) Cellulose in the cell walls of the Bangiophyceae (Rhodophyta). Science 207, 779–781

    Google Scholar 

  • Hawkes, M.W. (1978) A field, culture and ecological study of Porphyra gardneri, P. nereocystis, and P. thuretii (Rhodophyta, Bangiophycidae). Ph.D. dissertation, University of British Columbia, Vancouver, Canada

    Google Scholar 

  • Hepler, P.K. (1981) The structure of the endoplasmic reticulum revealed by osmium tetroxide-potassium ferricyanide staining. Eur. J. Cell Biol. 26, 102–110

    Google Scholar 

  • Kapraun, D.F., Freshwater, D.W. (1987) Karyological studies of five species of Porphyra (Bangiales, Rhodophyta) from the North Atlantic and Mediterranean. Phycologia 26, 82–87

    Google Scholar 

  • Kloareg, B., Quatrano, R.S. (1987) Isolation of protoplasts from zygotes of Fucus distichus (L.) Powell. Plant Sci. 50, 189–194

    Google Scholar 

  • Larkin, P.J. (1976) Purification and viability determinations of plant protoplasts. Planta 128, 213–216

    Google Scholar 

  • Ma, J.H., Miura, A. (1984) Observations of the nuclear division in the conchospores and their germlings in Porphyra yezoensis Ueda. Jap. J. Phycol. 32, 373–378

    Google Scholar 

  • McLachlan, J. (1973) Growth media — marine. In: Handbook of phycological methods: culture methods and growth measurements, pp. 25–51, Stein, J., ed. Cambridge University Press, London, UK

    Google Scholar 

  • Merrill, J.E., Waaland, J.R. (1988) Cultivation of Porphyra in Washington State. (Abstr.) J. Phycol. 24, Suppl., 5

    Google Scholar 

  • Mitman, G.G. (1988) A karyological study of Porphyra umbilicalis (L.) J. Agardh from Nova Scotia, Canada. In: Algal experimental systems in cell biological research, Am. Soc. for Cell Biol. Summer Res. Conf., June 25–29, Airlie House Conference Center, Airlie, Va., USA, Abstr. No. 40

    Google Scholar 

  • Mukai, L.S., Craigie, J.S., Brown, R.G. (1981) Chemical composition and structure of the cell walls of the conchocelis and thallus phases of Porphyra tenera (Rhodophyceae). J. Phycol. 17, 192–198

    Google Scholar 

  • Mumford, T.F., Jr., Cole, K. (1977) Chromosome numbers for fifteen species in the genus Porphyra (Bangiales, Rhodophyta) from the west coast of North America. Phycologia 16, 373–377

    Google Scholar 

  • Nakahara, H., Nakamura, Y. (1973) Parthenogenesis, apogamy and apospory in Alaria crassifolia (Laminariales). Mar. Biol. 18, 327–332

    Google Scholar 

  • Ohme, M., Miura, A. (1988) Tetrad analysis in conchospore germlings of Porphyra yezoensis (Rhodophyta, Bangiales). Plant Sci. 57, 135–140

    Google Scholar 

  • Polne-Fuller, M., Gibor, A. (1984) Developmental studies in Porphyra. I. Blade differentiation in Porphyra perforata as expressed by morphology, enzymatic digestion, and protoplast regeneration. J. Phycol. 20, 607–618

    Google Scholar 

  • Polne-Fuller, M., Biniaminov, M., Gibor, A. (1984) Vegetative propagation of Porphyra perforata. Hydrobiologia 116–117, 308–313

    Google Scholar 

  • Polne-Fuller, M., Saga, N., Gibor, N. (1986) Algal cell, callus, and tissue cultures and selection of algal strains. In: Algal biomass technologies: an interdisciplinary perspective, pp. 30–36, Barclay, W.R., McIntosh, R.P., eds. Beih. Nova Hedwigia No. 83. Cramer, Berlin, Germany

    Google Scholar 

  • Provasoli, L., Carlucci, A.F. (1974) Vitamins and growth regulators. In: Algal physiology and biochemistry, pp. 741–787, Stewart, W.D.P., ed. Bot. Monogr. No. 10. University of California Press, Berkeley, USA

    Google Scholar 

  • Pueschel, C.M. (1987) Absence of cap membranes as characteristic of pit plugs of some red algal orders. J. Phycol. 23, 150–156

    Google Scholar 

  • Saga, N., Sakai, Y. (1984) Isolation of protoplasts from Laminaria and Porphyra. Bull. Jap. Soc. Sci. Fish. 50, 1085

    Google Scholar 

  • Saga, N., Pome-Fuller, M., Gibor, A. (1986) Protoplasts from seaweeds: production and fusion. In: Algal biomass technologies: an interdisciplinary perspective, pp. 37–43, Barclay, W.R., McIntosh, R.P., eds. Beih. Nova Hedwigia No. 83. Cramer, Berlin, Germany

    Google Scholar 

  • Scott, J., Bosco, C., Schornstein, K., Thomas, J. (1980) Ultrastructure of cell division and reproductive differentiation of male plants in the Florideophyceae (Rhodophyta): cell division in Polysiphonia. J. Phycol. 16, 507–524

    Google Scholar 

  • Tang, Y. (1982) Isolation and culture of vegetative cells and protoplasts of Porphyra suborbiculata Kjellm. [Chin., Engl. summary.] J. Shandong Coll. Oceanology 12, 38–47

    Google Scholar 

  • van der Meer, J.P. (1988) Reproductive mutants of Gelidium sp. III Int. Phycol. Congr., Monash University, Melbourne, Australia, Aug. 14–20, Abstr., p. 43

    Google Scholar 

  • Waaland, S.D., Waaland, J.R. (1975) Analysis of cell elongation in red algae by fluorescent labelling. Planta 126, 127–138

    Google Scholar 

  • Woessner, J.W. (1977) The economic potential of harvesting and marketing an American Porphyra. (Abstr.) J. Phycol., 13, Suppl., 74

    Google Scholar 

  • Wynne, M.J., Loiseaux, S. (1976) Recent advances in life history studies of the Phaeophyta. Phycologia 15, 435–452

    Google Scholar 

Download references

Author information

Author notes

  1. Bruce A. Watson

    Present address: Department of Oenological Chemistry, Columbia Winery, 14030 NE 14th St., 98072, Woodinville, WA, USA

Authors and Affiliations

  1. Department of Botany, University of Washington, 98195, Seattle, WA, USA

    J. Robert Waaland, Leal G. Dickson & Bruce A. Watson

Authors
  1. J. Robert Waaland
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Leal G. Dickson
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Bruce A. Watson
    View author publications

    You can also search for this author in PubMed Google Scholar

Additional information

This paper is dedicated to the memory of the late Dr. Munenao Kurogi (1921–1988), Professor Emeritus of Hokkaido University

This research was supported by the Washington Sea Grant Program (National Oceanic and Atmospheric Administration). We thank Professor Y. Fujita (Nagasaki University, Japan), Professor S.-J. Wang (Shanghai University of Fisheries, P.R. China) and Dr. H. Kito (Seikai Regional Fisheries Research Laboratory, Nagasaki, Japan) for sharing their experience with Porphyra protoplast production with us. We thank J.S. Charleston for expert technical assistance in preparation of the electron-microscopy specimens. We also thank Dr. S.K. Herbert and John Carrier (Friday Harbor Laboratories) and Dr. John Merrill and D. Gillingham (American Sea Vegetable Co. and Applied Algal Research, Seattle) for collections of P. nereocystis.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Waaland, J.R., Dickson, L.G. & Watson, B.A. Protoplast isolation and regeneration in the marine red alga Porphyra nereocystis . Planta 181, 522–528 (1990). https://doi.org/10.1007/BF00193005

Download citation

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00193005

Key words

Search

Navigation