Multifunctional SEVA shuttle vectors for actinomycetes and Gram-negative bacteria

Area of Science:

• Microbiology
• Synthetic Biology
• Molecular Biology

Background:

• Actinomycetales, like Streptomyces, are valuable for producing secondary metabolites.
• Genetic engineering of Streptomyces is challenging due to limited genetic tools, hindering synthetic biology applications.

Purpose of the Study:

• To expand the genetic toolbox for Streptomyces and other actinomycetes.
• To develop novel shuttle vectors based on the SEVA architecture.

Main Methods:

• Construction of 23 novel SEVA shuttle vectors with dual origins of replication (Gram-negative and Streptomyces).
• Inclusion of vectors for integrative, low-copy, and medium-to-high-copy applications.
• Development of a new apramycin resistance selection marker.
• Testing vector functionality through heterologous expression of GFP and apigenin production in Streptomyces albus J1074.

Main Results:

• Successful development and characterization of 23 novel SEVA shuttle vectors.
• Demonstrated functionality via heterologous expression of GFP.
• Achieved heterologous production of the plant flavonoid apigenin.

Conclusions:

• The new SEVA vectors significantly enhance the genetic manipulation capabilities for Streptomyces species.
• These tools facilitate synthetic biology applications in actinomycetes.
• The expanded genetic toolbox will accelerate the use of Streptomyces as cell factories.