Cell Sorting-Directed Selection of Bacterial Cells in Bigger Sizes Analyzed by Imaging Flow Cytometry during...

Area of Science:

• Microbiology
• Synthetic Biology
• Evolutionary Biology

Background:

• Cell morphology is a key phenotypic trait for tracking adaptation and evolution.
• Quantitative techniques enable morphology tracking in large cell populations during experimental evolution.
• Directed evolution of novel cell morphologies has potential applications in synthetic biology, such as refining fermentation processes.

Purpose of the Study:

• To investigate the speed and feasibility of obtaining stable mutants with distinct morphologies using fluorescence-activated cell sorting (FACS)-directed experimental evolution.
• To explore the potential of combining FACS and imaging flow cytometry (IFC) for real-time tracking and selection of evolving bacterial populations.

Main Methods:

• Experimental evolution of E. coli populations using FACS for continuous sorting and culturing of cells with specific optical properties.
• Utilizing imaging flow cytometry (IFC) for real-time monitoring of population evolution.
• Genome sequencing to identify genetic mutations responsible for observed morphological changes.

Main Results:

• After ten rounds of FACS-directed evolution, a stable lineage of large E. coli cells was obtained.
• The large cell phenotype resulted from incomplete closure of the division ring.
• Genome sequencing identified a stop-gain mutation in the amiC gene, leading to a dysfunctional AmiC division protein.

Conclusions:

• FACS-directed experimental evolution is a rapid method for selecting and culturing bacteria with novel morphologies.
• The combination of FACS and IFC provides a powerful platform for real-time monitoring and directed evolution of microbial populations.
• This approach holds promise for generating new microbial phenotypes with potential applications in various fields, including synthetic biology.