Open AccessProtoplast preparation and regeneration from spores of the biocontrol fungus Pseudozyma flocculosa
Author(s) -
Cheng Yali,
Bélanger Richard R
Publication year - 2000
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2000.tb09300.x
Subject(s) - protoplast , spore , sucrose , biological pest control , regeneration (biology) , fungus , biology , microbiology and biotechnology , chemistry , biochemistry , botany
This paper describes a specific protocol for yielding and regenerating protoplasts from spores of the recently described powdery mildew biocontrol agent Pseudozyma flocculosa . With this protocol, a large quantity of protoplasts was obtained from β‐mercaptoethanol‐pretreated spores in 3‐day‐old cultures of P. flocculosa grown in YMPD. Enzymatic digestion was optimal with 0.5% Novozym 234 and 5% Glucanex prepared in 0.6 M KCl in 0.1 M citrate buffer. All liberated protoplasts fluoresced in the presence of fluorescein diacetate indicating that viability was nearly 100%. The regeneration rate was equally outstanding reaching 75% when 0.8 M sucrose was used as osmotic stabilizer in the regeneration medium. This protocol will find useful applications in genetic studies of this poorly characterized and understood biocontrol agent.