Reference : New disruption cassettes for rapid gene disruption and marker rescue in the yeast Yar...
Scientific journals : Article
Life sciences : Biochemistry, biophysics & molecular biology
Life sciences : Microbiology
New disruption cassettes for rapid gene disruption and marker rescue in the yeast Yarrowia lipolytica
Fickers, Patrick mailto [Université de Liège - ULg > > Centre d'ingénierie des protéines >]
Le Dall, M. T. [> > > >]
Gaillardin, C. [> > > >]
Thonart, Philippe mailto [Université de Liège - ULg > Gembloux Agro-Bio Tech > Gembloux Agro-Bio Tech - Biochimie et microbiologie industrielles >]
Nicaud, J. M. [> > > >]
Journal of Microbiological Methods
Elsevier Science Bv
Yes (verified by ORBi)
[en] gene disruption ; Cre-lox ; recombinase ; Yarrowia lipolytica
[en] Yarrowia lipolytica is one of the most extensively studied nonconventional yeasts. Unfortunately, few methods for gene disruption have been reported for this yeast, and all of them are time-consuming and laborious. The functional analysis of unknown genes requires powerful disruption methods. Here, we describe such a new method for rapid gene disruption in Y lipolytica. This knockout system combines SEP method and the Cre-lox recombination system, facilitating efficient marker rescue. Versatility was increased by using both auxotrophic markers like ylURA3 and ylLEU2, as well as the antibiotic resistance marker hph. The hph marker, which confers resistance to hygromycin-B, allows gene disruption in a strain lacking any conventional auxothrophic marker. The disruption cassette was shown to integrate at the correct locus at an average frequency of 45%. Upon expression of Cre recombinase, the marker was excised at a frequency of 98%, by recombination between the two lox sites. This new method for gene disruption is an ideal tool for the functional analysis of gene families, or for creating large-scale mutant collections in general. (C) 2003 Elsevier B.V. All rights reserved.

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