Miniaturized and automated adaptive laboratory evolution: Evolving Corynebacterium glutamicum towards an improved...

Area of Science:

• Biotechnology
• Microbial Engineering
• Synthetic Biology

Background:

• Adaptive Laboratory Evolution (ALE) is a key technique for untargeted microbial strain optimization.
• Efficient d-xylose utilization is crucial for developing sustainable biorefineries.
• Corynebacterium glutamicum is a versatile host for industrial biotechnology.

Purpose of the Study:

• To develop an automated and miniaturized ALE approach using microtiter plates.
• To improve the d-xylose utilization capabilities of Corynebacterium glutamicum.
• To identify genetic modifications responsible for enhanced d-xylose metabolism.

Main Methods:

• Automated, miniaturized Adaptive Laboratory Evolution (ALE) in microtiter plates.
• Cultivation of Corynebacterium glutamicum pEKEx3-xylXABCD_Cc on d-xylose.
• Bioreactor cultivation for stability assessment.
• Genome sequencing to identify mutations.

Main Results:

• Obtained an evolved strain, WMB2_evo, with a specific growth rate of 0.26h^-1 on d-xylose.
• Demonstrated stable growth of WMB2_evo in lab-scale bioreactors.
• Identified potential key mutations, including in the cg0196 gene (IolR regulator).

Conclusions:

• The automated ALE method effectively enhances microbial performance.
• The evolved strain WMB2_evo shows significant potential for biorefinery applications.
• Understanding genetic mutations provides insights for rational engineering of d-xylose utilization.