Skip to main content
SpringerLink
Log in

Protoplast isolation and regeneration from the potential economic brown alga Petalonia fascia (Ectocarpales, Phaeophyceae)

  • Published:
Journal of Applied Phycology Aims and scope Submit manuscript

Abstract

Petalonia fascia is a widespread brown alga with economic potential due to its use as raw or dried powder, and in the biomedical field. Protoplasts are living plant cells devoid of cell wall with a wide range of applications in basic and applied research, especially in crop improvement, and as a seedstock in seaweeds. Protoplasts have been previously isolated from P. fascia, but, their regeneration ability, an important prerequisite for protoplast applications, has not been explored. In this work, we report the protoplast isolation and successful regeneration from P. fascia using the commercially available cellulase “Onozuka” RS (1%) and alginate lyase (4 U mL−1). Protoplast production was enhanced under increased osmolarity (2512 mOsm L−1 H2O), with chelation pre-treatment, and short incubation time (4 h). Our protocol produced more than 14 times the number of protoplasts obtained using previously reported protocols. After 4 weeks in culture, protoplasts developed into prostrate and discoid and mixed thallus, as well as cell clumps. Blades mostly emerged from prostrate thalli. Antibiotics were not crucial for improving protoplast regeneration, and temperature did not affect the development of the morphological forms and blades. Our results show that high yields of protoplasts (107 protoplasts g−1 FW) with good regeneration ability can be obtained from P. fascia using a simple mixture of commercial enzymes. This represents the first report of successful protoplast regeneration in P. fascia.

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

Fig. 1
Fig. 2

Similar content being viewed by others

Data availability

The data that support the findings of this study are available from the corresponding author, Tae Oh Cho, upon reasonable request.

References

  • Abràmoff MD, Magalhães PJ, Ram SJ (2004) Image processing with ImageJ. Biophoton Int 11:36–42

    Google Scholar 

  • Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Avila-Peltroche J, Won BY (2020) Protoplast production from Sphacelaria fusca (Sphacelariales, Phaeophyceae) using commercial enzymes. J Mar Biosci Biotechnol 12:50–58

    Google Scholar 

  • Avila-Peltroche J, Won BY, Cho TO (2020) Optimization of protoplast isolation from the gametophytes of brown alga Undaria pinnatifida using response surface methodology. J Appl Phycol 32:2233–2244

    Article  CAS  Google Scholar 

  • Avila-Peltroche J, Won BY, Cho TO (2019) Protoplast isolation and regeneration from Hecatonema terminale (Ectocarpales, Phaeophyceae) using a simple mixture of commercial enzymes. J Appl Phycol 31:1873–1881

    Article  CAS  Google Scholar 

  • Benet H, Gall AE, Asensi A, Kloareg B (1997) Protoplast regeneration from gametophytes and sporophytes of some species in the order Laminariales (Phaeophyceae). Protoplasma 199:39–48

    Article  Google Scholar 

  • Boo SM (2010) Scytosiphonaceae, Petrospongiaceae. In: Kim HS, Boo SM (eds) Algal flora of Korea, volume 2, number 1. Heterokontophyta: Phaeophyceae: Ectocarpales. Marine brown algae I. National Institute of Biological Resources, Incheon

  • Burris KP, Dlugosz EM, Collins AG, Stewart CN, Lenaghan SC (2016) Development of a rapid, low-cost protoplast transfection system for switchgrass (Panicum virgatum L.). Plant Cell Rep 35:693–704

    Article  CAS  PubMed  Google Scholar 

  • Bustamante DE, Won BY, Cho TO (2016) The conspecificity of Pterosiphonia spinifera and P. arenosa (Rhodomelaceae, Ceramiales) inferred from morphological and molecular analyses. Algae 31:105–115

    Article  Google Scholar 

  • Butler DM, Ostgaard K, Boyen C, Evans LV, Jensen A, Kloareg B (1989) Isolation conditions for high yields of protoptasts from Laminaria saccharina and L. digitata (Phaeophyceae). J Exp Bot 40:1237–1246

    Article  CAS  Google Scholar 

  • Chen C-S, Shyu J-F (1994a) Isolation of protoplasts from four species of brown algae. Bot Bull Acad Sin 35:95–104

    Google Scholar 

  • Chen C-S, Shyu J-F (1994b) Regeneration of the protoplasts from the brown alga, Endarachne binghamiae (Phaeophyta; Punctariales, Scytosiphonaceae). Bot Bull Acad Sin 35:189–193

    Google Scholar 

  • Coelho SM, Scornet D, Rousvoal S, Peters N, Dartevelle L, Peters AF, Cock JM (2012) Isolation and regeneration of protoplast from Ectocarpus. Cold Spring Harbor Protoc. https://doi.org/10.1101/pdb.prot067959

  • Cribari-Neto F, Zeileis A (2010) Beta regression in R. J Stat Softw 34:1–24

    Article  Google Scholar 

  • Davey MR, Anthony P, Power JB, Lowe KC (2005) Plant protoplasts: status and biotechnological perspectives. Biotechnol Adv 23:131–171

    Article  CAS  PubMed  Google Scholar 

  • Elhai JD, Calhoun PS, Ford JD (2008) Statistical procedures for analysing mental health services data. Psychiatry Res 160:129–136

    Article  PubMed  Google Scholar 

  • Ferrari SLP, Cribari-Neto F (2004) Beta regression for modelling rates and proportions. J Appl Stat 31:799–815

    Article  Google Scholar 

  • Field A (2009) Discovering statistics using SPSS, 3rd edn. Sage, Los Angeles

    Google Scholar 

  • Fisher DD, Gibor A (1987) Production of protoplasts from the brown alga, Sargassum muticum (Yendo) Fensholt (Phaeophyta). Phycologia 26:488–495

    Article  Google Scholar 

  • Fox J, Weisberg S (2019) An R companion to applied regression, 3rd edn. Sage, Los Angeles https://socialsciences.mcmaster.ca/jfox/Books/Companion/

    Google Scholar 

  • Gupta V, Trivedi N, Simoni S, Reddy CRK (2018) Marine macroalgal nursery: a model for sustainable production of seedlings for large scale farming. Algal Res 31:463–468

    Article  Google Scholar 

  • Hothorn T, Bretz F, Westfall P (2008) Simultaneous inference in general parametric models. Biom J 50:346–363

    Article  PubMed  Google Scholar 

  • Hsiao SIC (1969) Life history and iodine nutrition of the marine brown alga, Petalonia fascia (O.F. Müll.) Kuntze. Can J Bot 47:1611–1616

    Article  CAS  Google Scholar 

  • Hsiao SIC (1970) Light and temperature effects on the growth, morphology, and reproduction of Petalonia fascia. Can J Bot 48:1359–1361

    Article  Google Scholar 

  • Huang L, Zhou J, Li X, Peng Q, Lu H, Du Y (2013) Characterization of a new alginate lyase from newly isolated Flavobacterium sp. S20. J Ind Microbiol Biotechnol 40:113–122

    Article  PubMed  Google Scholar 

  • Jackman S (2015) pscl: classes and methods for R developed in the Political Science Computational Laboratory, Department of Political Science. Stanford University, Stanford, California. R package version 1.4.9. URL http://pscl.stanford.edu/.

  • Kevekordes K, Beardall J, Clayton MN (1993) A novel method for extracting protoplasts from large brown algae. J Exp Bot 44:1587–1593

    Article  Google Scholar 

  • Kim T-H, Jung W-K (2019) R&D Trends of brown algae as potential candidates in biomedical application. J Mar Biosci Biotechnol 11:1–13

    CAS  Google Scholar 

  • Kloareg B, Polne-Fuller M, Gibor A (1989) Mass production of viable protoplasts from Macrocystis pyrifera. Plant Sci 62:105–112

    Article  Google Scholar 

  • Kogame K (1997) Sexual reproduction and life history of Petalonia fascia. Phycologia 36:389–394

    Article  Google Scholar 

  • Kogame K, Horiguchi T, Masuda M (1999) Phylogeny of the order Scytosiphonales (Phaeophyceae) based on DNA sequences of rbcL, partial rbcS, and partial LSU nrDNA. Phycologia 38:496–502

    Article  Google Scholar 

  • Lakens D (2013) Calculating and reporting effect sizes to facilitate cumulative science: a practical primer for t-tests and ANOVA. Front Psychol 4:863

    Article  PubMed  PubMed Central  Google Scholar 

  • Lee K-W, Cho J-H, Shin J-A (2003) A study on cultivation of Petalonia fascia (Scytosiphonales, Phaeophyta) by vegetative regeneration. Algae 18:333–339

    Article  Google Scholar 

  • Matsumoto K, Ichihara K, Shimada S (2014) Taxonomic reinvestigation of Petalonia (Phaeophyceae, Ectocarpales) in southeast of Honshu, Japan, with a description of Petalonia tenuis sp. nov. Phycologia 53:127–136

    Article  Google Scholar 

  • Matsumura W (1998) Efficient isolation and culture of viable protoplasts from Laminaria longissima Miyabe (Phaeophyceae). Bull Fish Sci Hokkaido Univ 49:85–90

    Google Scholar 

  • Mejjad M, Loiseaux-de-Goër S, Ducreux G (1992) Protoplast isolation, development, and regeneration in different strains of Pilayella littoralis (L.) Kjellm. (Phaeophyceae). Protoplasma 169:42–48

    Article  Google Scholar 

  • Peters G (2018) userfriendlyscience: quantitative analysis made accessible. https://doi.org/10.17605/osf.io/txequ. R package version 0.7.2: https://userfriendlyscience.com

  • Provasoli L (1968) Media and prospects for the cultivation of marine algae. In: Watanabe A, Hattori A (eds) Cultures and collections of algae. Proceedings of the U.S.–Japan Conference, Hakone, pp 63–75

  • R Core Team (2016). R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. https://www.R-project.org/. Accessed 3 Mar 2021

  • Reddy CRK, Gupta MK, Mantri VA, Bhavanath J (2008) Seaweed protoplast: status, biotechnological perspectives and needs. J Appl Phycol 20:619–632

    Article  CAS  Google Scholar 

  • Salmeán AA, Duffieux D, Harholt J, Qin F, Michel G, Czjzek M, Willats WGT, Hervé C (2017) Insoluble (1→3), (1→4)-β-D-glucan is a component of cell walls in brown algae (Phaeophyceae) and is masked by alginates in tissues. Sci Rep 7:2880

    Article  PubMed  PubMed Central  Google Scholar 

  • Sellke T, Bayarri MJ, Berger JO (2001) Calibration of p values for testing precise null hypotheses. Am Stat 55:62–71

    Article  Google Scholar 

  • Smithson M, Verkuilen J (2006) A better lemon squeezer? Maximum-likelihood regression with Beta-distributed dependent variables. Psychol Methods 11:54–71

    Article  PubMed  Google Scholar 

  • Sullivan GM, Feinn R (2012) Using effect size-or why the P value is not enough. J Grad Med Educ 4:279–282

    Article  PubMed  PubMed Central  Google Scholar 

  • Thibault J-F, Rouau X (1990) Studies on enzymic hydrolysis of polysaccharides in Sugar beet pulp. Carbohydr Polym 13:1–16

    Article  CAS  Google Scholar 

  • Venables WN, Ripley BD (2002) Modern applied statistics with S, 4th edn. Springer, New York

    Book  Google Scholar 

  • Xiaoke H, Xiaolu J, Huashi G (2003) Isolation of protoplast from Undaria pinnatifida by alginate lyase digestion. J Ocean Univ China 2:58–61

    Article  Google Scholar 

Download references

Funding

This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (2019R1F1A1060346 and NRF-2021R1I1A2059577), by the Ministry of Ocean and Fisheris (MarineBiotics Project, 20210469), and by the National Marine Biodiversity Institute of Korea (2021M01100) to Tae Oh Cho. This research was also supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-2021R1I1A1A01051909) to Boo Yeon Won.

Author information

Authors and Affiliations

  1. Department of Life Science, Chosun University, Gwangju, 61452, Korea

    Jose Avila-Peltroche, Boo Yeon Won & Tae Oh Cho

  2. Department of Integrative Biological Sciences & BK21 FOUR Educational Research Group for Age-associated Disorder Control Technology, Chosun University, Gwangju, 61452, Korea

    Jose Avila-Peltroche & Tae Oh Cho

Authors
  1. Jose Avila-Peltroche
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Boo Yeon Won
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tae Oh Cho
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

All authors contributed to the study conception and design. Material preparation, data collection, and analysis were performed by Jose Avila-Peltroche. The first draft of the manuscript was written by Jose Avila-Peltroche and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Tae Oh Cho.

Ethics declarations

Conflict of interest

The authors declare no competing interests.

Additional information

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Avila-Peltroche, J., Won, B.Y. & Cho, T.O. Protoplast isolation and regeneration from the potential economic brown alga Petalonia fascia (Ectocarpales, Phaeophyceae). J Appl Phycol 34, 543–550 (2022). https://doi.org/10.1007/s10811-021-02648-0

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10811-021-02648-0

Keywords

Search

Navigation