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Protoplast isolation and regeneration from Gracilaria changii (Gracilariales, Rhodophyta)

Journal of Applied Phycology volume 20, pages 641–651 (2008)Cite this article

Abstract

The tropical agarophyte Gracilaria changii has been much researched and documented by the Algae Research Laboratory, University of Malaya, especially with regards to its potential as a seaweed bioreactor for valuable compounds. Protoplast regeneration of this seaweed was developed following the optimization of protoplast isolation protocol. Effect of the concentration and combination of isolating enzymes, incubation period, temperature, enzyme solution pH, tissue source on the protoplast yields were used to optimize the isolation protocol. The enzyme mixture with 4% w/v cellulase Onozuka R-10, 2% w/v macerozyme R-10 and 1 unit mL-1 agarase was found to produce the highest yield of protoplast at 28°C and 3 h incubation period. Thallus tips gave higher yields of protoplasts than middle segments. Freshly isolated G. changii protoplasts were cultured in MES medium. Regeneration of protoplast cell walls after 24 h was confirmed by calcofluor white M2R staining under UV fluorescence microscopy. The protoplasts with regenerated cell walls then underwent a series of cell division to produce callus-like cell masses in MES medium. Following this, juvenile plants of G. changii were obtained.

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Acknowledgments

This research was funded by the Ministry of Science, Technology and Innovation, Malaysia (Grant No: 39-02-03-9002/Oracle 8301902; 12-02-03-2028/Oracle 8422028), University of Malaya Short-term Research Grant No. F0140/2002D, F0133/2004A, F0233/2004D and P0114/2006A. Ms Yeong conducted this research under a PASCA scholarship from the University of Malaya. The authors gratefully acknowledge the following for their advice and kind assistance: Professor He Peimin, Dr. Gan Sook Yee, Dr. Tan Eng Lai, Professor Qin Song and Dr. Jiang Peng.

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Authors and Affiliations

  1. Institute of Postgraduate Studies, University of Malaya, 50603, Kuala Lumpur, Malaysia

    Hui-Yin Yeong

  2. Institute of Biological Sciences, University of Malaya, 50603, Kuala Lumpur, Malaysia

    Norzulaani Khalid & Siew-Moi Phang

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  1. Hui-Yin Yeong
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  2. Norzulaani Khalid
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  3. Siew-Moi Phang
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Correspondence to Siew-Moi Phang.

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Yeong, HY., Khalid, N. & Phang, SM. Protoplast isolation and regeneration from Gracilaria changii (Gracilariales, Rhodophyta). J Appl Phycol 20, 641–651 (2008). https://doi.org/10.1007/s10811-007-9249-5

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  • DOI: https://doi.org/10.1007/s10811-007-9249-5

Keywords

  • Callus
  • Cell wall degrading enzymes
  • Gracilaria changii
  • Protoplast isolation
  • Protoplast regeneration