ArticleMicrobial biotechnology2020
Protocols for RecET-based markerless gene knockout and integration to express heterologous biosynthetic gene clusters in Pseudomonas putida.
Article in Microbial biotechnology, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 12 papers.
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Who cites it
12 citing papers in PubMed.
- Identification of proteins influencing CRISPR-associated transposases for enhanced genome editing.Science advances · 2026Article
- Enhanced production of microbial levulinic acid through deletion of the levulinic acid transcriptional regulator (lvaR) in engineered Pseudomonas putida KT2440.Bioprocess and biosystems engineering · 2025Article
- Modification of Glucose Metabolic Pathway to Enhance Polyhydroxyalkanoate Synthesis inCurrent issues in molecular biology · 2024Article
- Computational analysis into the potential of azo dyes as a feedstock for actinorhodin biosynthesis in Pseudomonas putida.PloS one · 2024Article
- Microbial antibiotics take the lead in the fight against plant pathogens.Microbial biotechnology · 2023Article
- The modular pYT vector series employed for chromosomal gene integration and expression to produce carbazoles and glycolipids inFEMS microbes · 2023Article
- Industrial biotechnology of Pseudomonas putida: advances and prospects.Applied microbiology and biotechnology · 2020Review
- Synthetic control of plasmid replication enables target- and self-curing of vectors and expedites genome engineering ofMetabolic engineering communications · 2020Article
- Protocols for RecET-based markerless gene knockout and integration to express heterologous biosynthetic gene clusters in Pseudomonas putida.Microbial biotechnology · 2020Article
- Accelerated genome engineering of Pseudomonas putida by I-SceI-mediated recombination and CRISPR-Cas9 counterselection.Microbial biotechnology · 2020Article
- Unraveling Plant Natural Chemical Diversity for Drug Discovery Purposes.Frontiers in pharmacology · 2020Review
- Bacterial Genetic Engineering by Means of Recombineering for Reverse Genetics.Frontiers in microbiology · 2020Review
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2 authors.
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Abstract
Pseudomonas putida has emerged as a promising host for the production of chemicals and materials thanks to its metabolic versatility and cellular robustness. In particular, P. putida KT2440 has been officially classified as a generally recognized as safe (GRAS) strain, which makes it suitable for the production of compounds that humans directly consume, including secondary metabolites of high importance. Although various tools and strategies have been developed to facilitate metabolic engineering of P. putida, modification of large genes/clusters essential for heterologous expression of natural products with large biosynthetic gene clusters (BGCs) has not been straightforward. Recently, we reported a RecET-based markerless recombineering system for engineering P. putida and demonstrated deletion of multiple regions as large as 101.7 kb throughout the chromosome by single rounds of recombineering. In addition, development of a donor plasmid system allowed successful markerless integration of heterologous BGCs to P. putida chromosome using the recombineering system with examples of - but not limited to - integrating multiple heterologous BGCs as large as 7.4 kb to the chromosome of P. putida KT2440. In response to the increasing interest in our markerless recombineering system, here we provide detailed protocols for markerless gene knockout and integration for the genome engineering of P. putida and related species of high industrial importance.
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