Preparation of protoplasts from the carrageenophyte Gigartina corymbifera (Kutz.) J. Ag. (Rhodophyta)
1990
Gross, W.
Protoplasts were isolated with high yield from the carrageenophyte Gigartina corymbifera (Kutz.) J. Ag. by using the enzyme carrageenase in combination with cell wall-digesting enzymes. The enzyme mixture consisted of 5 U carrageenase.ml-1, 2% cellulase, 2% Macerozyme R-10, and 0.2% Pectolyase Y-23 dissolved in 60% seawater containing 0.7 M mannitol, 5 mM CaCl2, and 40 mM Tris-HCl, pH 7. Carrageenase was prepared from cultures of the marine bacterium Pseudomonas carrageenovora. Protoplasts from G. corymbifera were spherical, dark red-colored and very uniform in size (approximately 17 micrometer); they originated solely from the epidermal tissue and > 95% of the protoplasts were viable. Freshly prepared protoplasts lack cell walls and polysaccharide sheaths, as demonstrated by electron microscopy and specific staining methods. Spontaneous cell fusion was observed on several occasions. These protoplasts could serve as a useful tool in crop improvement of this important carrageenan-producing alga.
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