An efficient genetic transformation method for glycerol producer Candida glycerinogenes

Transformation techniques generally require development before genetic and molecular studies of industrial yeast strains can commence. Candida glycerinogenes WL2002-5 has been used for industrial-scale glycerol production but has proven difficult to transform for molecular studies following previous...

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Bibliographic Details
Published in:Microbiological research 2008-09, Vol.163 (5), p.531-537
Main Authors: Chen, Xianzhong, Fang, Huiying, Rao, Zhiming, Shen, Wei, Zhuge, Bin, Wang, Zhengxiang, Zhuge, Jian
Format: Article
Language:English
Subjects:
Acetates - pharmacology
Anti-Bacterial Agents - pharmacology
Candida - drug effects
Candida - genetics
Candida - metabolism
Candida glycerinogenes
Dithiothreitol - pharmacology
Electroporation
Gene Transfer Techniques
Genome, Fungal
Glycerol - metabolism
Glycerol production
Industrial Microbiology
Plasmids - genetics
Transformation
Transformation, Genetic
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Summary:Transformation techniques generally require development before genetic and molecular studies of industrial yeast strains can commence. Candida glycerinogenes WL2002-5 has been used for industrial-scale glycerol production but has proven difficult to transform for molecular studies following previously published procedures. In the present study, phleomycin was used as drug-resistance marker based on perturbing plasma membranes and the lower minimum inhibitory concentration for C. glycerinogenes. We developed an efficient method for transformation of C. glycerinogenes, and parameters involved in transformation efficiency were optimized. Pretreatment of yeast cells with either dithiothreitol or lithium acetate enhanced the frequency of transformation markedly by electroporation. Significantly higher transformation efficiency was obtained when the electroporated cells were pretreated with phleomycin before plating. With this method, a maximal transformation frequency of 394±50 transformants/μg plasmid DNA was obtained. The transformation method will foster genetic manipulation of this industrial strain.
ISSN:0944-5013
1618-0623
DOI:10.1016/j.micres.2008.05.003