Editing the Serratia proteamaculans Genome Using the Allelic Exchange Method

Area of Science:

• Microbiology
• Molecular Biology
• Bacterial Genetics

Background:

• Serratia proteamaculans exhibits diverse ecological roles, including opportunistic pathogenicity and symbiosis, making it a valuable model for studying interspecific interactions.
• Understanding gene function in S. proteamaculans is crucial, but requires adapted genome editing tools due to species-specific characteristics.

Purpose of the Study:

• To adapt and validate an allelic exchange-based genome editing method for Serratia proteamaculans.
• To investigate the biological role of protealysin through targeted genetic modification.

Main Methods:

• Adaptation of homologous recombination via the allelic exchange method using the suicidal vector pRE118.
• Delivery of recombination plasmids to S. proteamaculans via conjugation using an auxotrophic Escherichia coli ST18 strain.
• Generation of genetic modifications without the need for inserting selective markers into the S. proteamaculans genome.

Main Results:

• Successful adaptation of the allelic exchange method for genome editing in S. proteamaculans.
• Achieved a selection strategy yielding 30%-40% of clones with the target mutation at the final stage.
• Demonstrated the method's validity for creating knockouts, knockins, and point mutations.

Conclusions:

• The developed genome editing method is efficient and versatile for S. proteamaculans research.
• This approach enables functional genomics studies without requiring selective markers, simplifying genetic manipulation.
• The method is adaptable for genome editing in other bacterial species, broadening its applicability.