A molecular genetic toolbox for Yarrowia lipolytica

Area of Science:

• Biotechnology
• Synthetic Biology
• Microbial Engineering

Background:

• Yarrowia lipolytica is a key ascomycete yeast for protein secretion and lipid accumulation.
• Existing genetic tools for Y. lipolytica vary in effectiveness across different strains.
• A comprehensively sequenced strain is crucial for advanced genetic manipulation.

Purpose of the Study:

• To develop novel genetic and molecular tools for Yarrowia lipolytica.
• To expand the capabilities of Y. lipolytica for research and industrial bioengineering.
• To create a foundational resource for Yarrowia cell biology and metabolism studies.

Main Methods:

• Generation of isogenic auxotrophic strains with reduced non-homologous end joining for precise DNA integration.
• Genome sequencing, assembly, and annotation to identify novel genes.
• Construction of plasmids with optimized superfolder GFP for overexpression and fluorescent tagging.
• Development of the Yarrowia lipolytica Cell Atlas with fluorescently tagged organelles.

Main Results:

• Established a set of isogenic auxotrophic Y. lipolytica strains.
• Uncovered previously unidentified genes within the Y. lipolytica genome.
• Created Y. lipolytica strains with endogenous fluorescently tagged organelles.
• Successfully built the Yarrowia lipolytica Cell Atlas for live cell imaging.

Conclusions:

• The developed tools facilitate live assessment of organelle-specific protein expression and localization.
• These resources are valuable for studying cell biology and metabolism in Y. lipolytica.
• This work supports the advancement of biofuels and natural product development using Y. lipolytica.