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High-throughput transformation method for Yarrowia lipolytica mutant library screening.

Christophe Leplat¹, Jean-Marc Nicaud¹, Tristan Rossignol²

¹INRA, UMR1319 Micalis, F-78352 Jouy-en-Josas, France AgroParisTech, UMR Micalis, F-78352 Jouy-en-Josas, France.

FEMS Yeast Research

|June 24, 2015

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Researchers developed a rapid, high-throughput method for genetically engineering Yarrowia lipolytica yeast. This new system enables large-scale screening of overexpression mutants, accelerating biotechnological advancements.

Area of Science:

Biotechnology
Microbiology
Synthetic Biology

Background:

The oleaginous yeast Yarrowia lipolytica is crucial for biotechnology and undergoes extensive genetic modification.
Developing large mutant collections for high-throughput screening is essential for future advancements.

Purpose of the Study:

To establish a rapid and efficient method for high-throughput transformation of Y. lipolytica.
To create plasmids and strains for large-scale screening of overexpression mutant strains.

Main Methods:

Adaptation of Gateway® vectors for specific locus integration in Y. lipolytica.
Development of a platform strain with constitutive fluorescent protein expression for growth monitoring.
Generation of alkaline extracellular protease (AEP) overexpression mutants in 96-well plates.

Keywords:

Gateway®XPR2 Yarrowia lipolytica overexpression transformation method, mutant screening

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Main Results:

A high-throughput transformation system for Y. lipolytica was successfully established.
A collection of AEP overexpression mutants was generated efficiently in a single experiment.
The platform strain facilitated screening in non-translucid media using a fluorescent growth reporter.

Conclusions:

The developed platform significantly enhances the capacity for genetic engineering and functional genomic analysis in Y. lipolytica.
This method accelerates the discovery and optimization of yeast strains for biotechnological applications.
The system provides a valuable tool for generating and screening large mutant libraries for industrial yeast development.