Dan Sun, Hui-Hui Li, Jing Wu, Jie Wu, Wei-Qiang Lin, Ru-Li He, Dong-Feng Liu, Wen-Wei Li
{"title":"Antibiotics-Free Steady Bioproduction of Valuable Chemicals from Organic Wastes by Engineered <i>Vibrio natriegens</i> through Targeted Gene Integration.","authors":"Dan Sun, Hui-Hui Li, Jing Wu, Jie Wu, Wei-Qiang Lin, Ru-Li He, Dong-Feng Liu, Wen-Wei Li","doi":"10.1021/acssynbio.4c00669","DOIUrl":null,"url":null,"abstract":"<p><p>Bioproduction of chemicals by using engineered bacteria is promising for a circular economy but challenged the instability of the introduced plasmid by conventional methods. Here, we developed a two-plasmid INTEGRET system to reliably integrate the targeted gene into the <i>Vibrio natriegens</i> genome, making it a powerful strain for efficient and steady bioproduction without requiring antibiotic addition. The INTEGRET system allows for gene insertion at over 75% inserting efficiency and flexibly controllable gene dosages. Additionally, simultaneous gene insertion at four genomic sites was achieved at 54.3% success rate while maintaining stable inheritance of exogenous sequences across multiple generations. The engineered strain could efficiently synthesize PHB from the fermentation of diverse organic wastes, with an efficiency comparable to those with overexpressed plasmid. When the mixture of seawater and molasses was used as the feedstock, it achieved a high PHB yield of 39.41 wt %. An extended application of the INTEGRET system for imparting the riboflavin production ability to the bacterium was also demonstrated. Our work presents a reliable and efficient genomic editing tool to facilitate the development of sustainable and environmentally benign biological platforms for converting biomass wastes into valuable chemicals.</p>","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":"4233-4244"},"PeriodicalIF":3.7000,"publicationDate":"2024-12-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"ACS Synthetic Biology","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1021/acssynbio.4c00669","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2024/12/3 0:00:00","PubModel":"Epub","JCR":"Q1","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
引用次数: 0
Abstract
Bioproduction of chemicals by using engineered bacteria is promising for a circular economy but challenged the instability of the introduced plasmid by conventional methods. Here, we developed a two-plasmid INTEGRET system to reliably integrate the targeted gene into the Vibrio natriegens genome, making it a powerful strain for efficient and steady bioproduction without requiring antibiotic addition. The INTEGRET system allows for gene insertion at over 75% inserting efficiency and flexibly controllable gene dosages. Additionally, simultaneous gene insertion at four genomic sites was achieved at 54.3% success rate while maintaining stable inheritance of exogenous sequences across multiple generations. The engineered strain could efficiently synthesize PHB from the fermentation of diverse organic wastes, with an efficiency comparable to those with overexpressed plasmid. When the mixture of seawater and molasses was used as the feedstock, it achieved a high PHB yield of 39.41 wt %. An extended application of the INTEGRET system for imparting the riboflavin production ability to the bacterium was also demonstrated. Our work presents a reliable and efficient genomic editing tool to facilitate the development of sustainable and environmentally benign biological platforms for converting biomass wastes into valuable chemicals.
期刊介绍:
The journal is particularly interested in studies on the design and synthesis of new genetic circuits and gene products; computational methods in the design of systems; and integrative applied approaches to understanding disease and metabolism.
Topics may include, but are not limited to:
Design and optimization of genetic systems
Genetic circuit design and their principles for their organization into programs
Computational methods to aid the design of genetic systems
Experimental methods to quantify genetic parts, circuits, and metabolic fluxes
Genetic parts libraries: their creation, analysis, and ontological representation
Protein engineering including computational design
Metabolic engineering and cellular manufacturing, including biomass conversion
Natural product access, engineering, and production
Creative and innovative applications of cellular programming
Medical applications, tissue engineering, and the programming of therapeutic cells
Minimal cell design and construction
Genomics and genome replacement strategies
Viral engineering
Automated and robotic assembly platforms for synthetic biology
DNA synthesis methodologies
Metagenomics and synthetic metagenomic analysis
Bioinformatics applied to gene discovery, chemoinformatics, and pathway construction
Gene optimization
Methods for genome-scale measurements of transcription and metabolomics
Systems biology and methods to integrate multiple data sources
in vitro and cell-free synthetic biology and molecular programming
Nucleic acid engineering.