Protoplast isolation from Dictyopteris pacifica and Scytosiphon lomentaria, using a simple commercial enzyme preparation

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

doi:10.1186/s43141-021-00226-y

Jose Avila-Peltroche, Boo Yeon Won + Show 1 more

Open Access

https://doi.org/10.1186/s43141-021-00226-y

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Abstract

BackgroundProtoplasts (i.e., naked plant cells) can be used for in vitro manipulations and genetic improvement in cultivars with economic value. During the last decade, protoplast research in economic brown algae has been scarce, and it is usually hampered by the use of non-commercial enzymes or crude extracts for isolating protoplasts. Dictyopteris pacifica is part of a brown algal genus well known by its wide chemical diversity and biological properties. Scytosiphon lomentaria is an edible brown seaweed with antioxidant, antitumor, and antiviral properties. So far, there are no protoplast isolation protocols using commercial enzymes for these two economic brown algae. In this study, we obtained protoplasts from cultured samples of D. pacifica and S. lomentaria using commercially available enzymes. Additionally, we investigated the effects of Driselase inclusion and Ca-chelation pre-treatment on protoplast yields in order to optimize the conditions for protoplast preparations. ResultsProtoplasts were isolated from Dictyopteris pacifica and Scytosiphon lomentaria using the commercially available Cellulase Onozuka RS (1%) and Alginate lyase (4 U mL−1), and short incubation time (4 h). Driselase did not show significant effects on protoplast production in both species. Ca-chelation pre-treatment only increased the number of protoplasts in D. pacifica. Under optimal conditions, the protoplast yields from D. pacifica and S. lomentaria were 4.83 ± 2.08 and 74.64 ± 32.49 × 106 protoplasts g−1 fresh weight, respectively. The values obtained for S. lomentaria were 2–3 orders of magnitude higher than previously reported. ConclusionsOur results show that high protoplast yields can be obtained from D. pacifica and S. lomentaria using a simple mixture of commercial enzymes (Cellulase RS and Alginate lyase) and short incubation time (4 h). This work also represents the first report of protoplast isolation in D. pacifica. The method proposed here can help to expand protoplast technology in more brown algal species.

Highlights

Protoplasts can be used for in vitro manipulations and genetic improvement in cultivars with economic value
This research aims to develop a protocol for protoplast production from Dictyopteris pacifica and Scytosiphon lomentaria as an effort to expand protoplast technology in commercial brown algae
The thalli did not show marked constrictions as reported by Boo [18], molecular analysis using a rbcL region (1342-bp) (MW715818) of our strain was matched with 99.85% to field samples of S. lomentaria from Korea and Japan [39]

Summary

Introduction

Protoplasts (i.e., naked plant cells) can be used for in vitro manipulations and genetic improvement in cultivars with economic value. Protoplast research in economic brown algae has been scarce, and it is usually hampered by the use of non-commercial enzymes or crude extracts for isolating protoplasts. There are no protoplast isolation protocols using commercial enzymes for these two economic brown algae. Protoplasts are cells whose cell walls have been removed mostly by enzymatic methods They are usually produced from plant cells, protoplasts can be obtained from bacteria [1] and fungi [2]. Most protocols rely on non-commercial enzymes or crude extracts for protoplast isolation, which make them expensive, time consuming and low reproducible [6, 7]

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