Rapid evolution of novel traits in microorganisms

Area of Science:

• Microbial Biotechnology
• Synthetic Biology
• Genetic Engineering

Background:

• Microbial biotransformations are limited by low tolerance to industrial conditions like high metabolite and solvent concentrations.
• Strain improvement often relies on random mutagenesis and selection, which can be slow.
• Mutator strains accelerate evolution but require restoring normal mutation rates post-selection to avoid accumulating errors.

Purpose of the Study:

• To develop a method for rapid microbial strain improvement.
• To accelerate directed evolution for enhanced industrial biotransformation capabilities.
• To enable temporary, controlled increases in mutation frequency for trait development.

Main Methods:

• Construction of temperature-sensitive plasmids to temporarily increase host mutation frequency (20- to 4,000-fold).
• Application of these mutator plasmids under selection pressure for directed evolution.
• Curing of plasmids to restore normal mutation rates in evolved strains.
• Testing the method on bacterial strains to improve tolerance to dimethylformamide (DMF).

Main Results:

• Successfully increased the tolerance of three bacterial strains to dimethylformamide by 10 to 20 g/liter within two transfers.
• Demonstrated rapid acquisition of new, complex traits in microorganisms.
• Showcased the ability to revert strains to a low mutation rate after selection by plasmid curing.

Conclusions:

• The developed temperature-sensitive plasmid system offers an efficient in vivo mutagenesis approach for rapid strain improvement.
• This method accelerates the evolution of microorganisms for enhanced industrial applications.
• It provides a controllable way to boost mutation rates for trait development and then stabilize the improved strains.