Application of an Alginate Lyase from Alteromonas sp. for Isolation of Protoplasts from a Brown Algae Laminaria japonica.

Abstract

An alginate lyase from Alteromonas sp H-4 was used for the preparation of protoplast from young sporophytes of Laminaria japonica. Optimum conditions for the isolation of protoplasts were as follows: a) fine pieces of the sporophytes chopped by a razor were incubated in an enzyme mixture for 3 to 5h at 15° with shaking, b) the composition of the enzyme mixture was purified alginate lyase (30U/ml) with cellulase Onozuka R-10 (1.5%) in a hypertonic solution (0.7M man nitol, 25mM MgCl2, and 5mM HEPES in 50% seawater pH7.8). Yields of protoplast were 106- 107 cells/g (sporophytes) and protoplast viability was more than 75% Protoplasts were browngreenish and spherical with a diameter of 10-25µm. The purity of alginate lyase from the strain H-4 scarcely influenced the yield of protoplast, but purified enzyme produced the highest viability of cells. © 1993, The Japanese Society of Fisheries Science. All rights reserved.