Morphogenetic responses from protoplasts and tissue culture of Laminaria digitata (Linnaeus) J. V. Lamouroux (Laminariales, Phaeophyta): callus and thalloid-like structures regeneration

摘要

The regeneration of meristematic tissues from sporophytes of Laminaria digitata was studied by protoplast and tissue culture. Sequential treatment of explants in sterile seawater with 1% Betadine for 5 min, 1% commercial bleach for 1-2 min and 2% antibiotic treatment supplemented with 1 oM GeO overnight enabled viable explants as high as 55%. Different morphogenetic responses were observed from tissue culture on media supplemented with plant growth regulators alone or in combination, mainly filamentous calluses up to 50% according to the media. Dark green compact calluses were observed on two combinations: 4 oM Pi + 2 oM N-(2-chloro-4-pyridyl)-N'-phenylurea (CPPU) and 0.04 oM Pi + 0.44 oM 6-benzylaminopurine. Thalloid-like structures comparable to adventitious buds were regenerated on medium supplemented with 4 oM Pi + 0.45 oM zeatin but at low frequency suggesting a strong genotypic effect. Friable calluses were developed from protoplasts in enriched medium with polyamines and containing 0.40 oM CPPU + 0.45 oM 2,4-dichlorophenoxyacetic acid. In order to produce protoplasts, a one-step enzymatic protocol was developed and yields reached 22 x 10e protoplasts per gram of fresh weight.