Non-enzymatic isolation of somatic cells fromKappaphycusspp. andEucheuma denticulatum(Solieriaceae, Rhodophyta)

Abstract

Cell culture technology is immensely useful in somatic hybridization, induction of mutations, cloning of specific isolates and maintenance of strains of defined genotypes. However, its application in strain improvement of some tropical red macroalgae has been limited due to the difficulty of isolating viable cells from their complex intercellular matrices. A simple, non-enzymatic technique of isolating somatic cells was developed for Kappaphycus spp. and Echeuma denticulatum. Surface-sterilized tissues (0.1 g fresh weight, 2.0 mm thick discs) from subcortical and medullary layers were treated with 3% NaOH, 3% KOH, or hydrogen peroxide in phosphate buffer solution (PBH 2 O 2 ). Tissues of K. alvarezii treated with PBH 2 O 2 softened after 5 h of treatment and completely dissociated after 12 h. Viable cell counts (VCC), determined through staining with Evan's blue, were significantly higher (2.4 × 10 5 cells g −1 fresh weight tissue) in K. alvarezii ('tambalang' strain) treated with PBH 2 O 2 compared with tissues treated with carrageenase from a marine bacterium.