Restructured Lactococcus lactis strains with emergent properties constructed by a novel highly efficient screening...

Area of Science:

• Microbiology
• Synthetic Biology
• Biotechnology

Background:

• Previous genome reduction of Lactococcus lactis NZ9000 yielded a promising protein production host.
• The traditional gene deletion process for genome reduction is time-consuming and labor-intensive.

Purpose of the Study:

• To develop a convenient and efficient method for large-scale genome reduction in L. lactis NZ9000.
• To streamline the generation of restructured L. lactis strains for enhanced industrial applications.

Main Methods:

• Construction of plasmid pNZ5417 with a P_nisZ-lacZ visually selectable marker.
• Stepwise deletion of two large nonessential DNA regions (L-4A and L-5A) in L. lactis 9k-3.

Main Results:

• The developed plasmid enabled more efficient and convenient screening of gene deletion mutants.
• Deletion of 1.25% of the chromosome resulted in the L. lactis 9k-5A mutant.
• The L. lactis 9k-5A mutant exhibited improved growth, transformability, carbon metabolism, and amino acid biosynthesis compared to the parent strain.

Conclusions:

• A convenient and efficient system for large-scale genome deletion in L. lactis was established using a visually selectable marker.
• This method facilitates rapid genome streamlining and the generation of engineered L. lactis cell factories.