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Automated Construction of a Yeast-Based Multigene Library via Homologous Recombination in a Biofoundry Workflow
Min-Jun Seong1,2, Ye Rin Yoon1, Kil Koang Kwon1,2
1Synthetic Biology Research Center and the K-Biofoundry, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea.
ACS Synthetic Biology
|May 7, 2025
Summary
This study introduces an automated one-step method for multigene assembly using in vivo homologous recombination, streamlining the creation of expression-tunable libraries for synthetic biology applications.
Area of Science:
- Synthetic Biology
- Molecular Biology
- Genetic Engineering
Background:
- Traditional cloning techniques limit efficient metabolic pathway construction.
- Existing in vitro gene assembly methods are complex and labor-intensive.
- A need exists for simplified, high-throughput gene assembly strategies.
Purpose of the Study:
- To develop an automated, one-step multigene assembly method.
- To create an expression-tunable library using in vivo homologous recombination.
- To enhance efficiency and throughput in genetic engineering.
Main Methods:
- Developed an automated one-step multigene assembly protocol.
- Utilized in vivo homologous recombination for gene assembly.
- Optimized a shuttle vector to improve recombination efficiency.
- Scaled the method for high-throughput applications.
Main Results:
- Successfully constructed an expression-tunable multigene library.
- Demonstrated the method's efficiency and automation capabilities.
- Validated the approach within a biofoundry setting.
Conclusions:
- The developed method offers a versatile strategy for genetic engineering.
- Enables parallel and high-throughput construction of gene libraries.
- Advances synthetic biology by simplifying complex genetic constructions.
