Development of a Transformation System for Nitratireductor sp
Hiroto Maeda1, Yuto Hirata1, Hirokazu Takahashi1
1Graduate School of Integrated Sciences for Life, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima, Hiroshima, 739-8530, Japan.
Marine Biotechnology (New York, N.Y.)
|February 2, 2023
Summary
Researchers developed a gene transfer system for Nitratireductor sp. OM-1, confirming the thioesterase (te) gene
Area of Science:
- Microbiology
- Synthetic Biology
- Biotechnology
Background:
- Nitratireductor sp. OM-1 accumulates butenoic acid, a valuable short-chain unsaturated carboxylic acid.
- The thioesterase (te) gene was previously predicted as a candidate for butenoic acid biosynthesis.
- A functional gene transfer system was lacking for Nitratireductor sp. OM-1.
Purpose of the Study:
- To establish a gene transfer system for Nitratireductor sp. OM-1.
- To confirm the role of the thioesterase (te) gene in butenoic acid production.
- To optimize conditions for electroporation-mediated transformation.
Main Methods:
- Electroporation was employed as the gene transfer method.
- A broad-host-range plasmid, pRK415, was utilized for constructing expression vectors.
- The thioesterase (te) gene was inserted into pRK415 to create pRK415-te.
Main Results:
- Optimized electroporation conditions yielded a maximum transformation efficiency of 7.9 × 10⁴ colonies/µg DNA at 22.5 kV/cm.
- The constructed pRK415-te vector was successfully transferred into Nitratireductor sp. OM-1.
- Recombinant OM-1 strains produced butenoic acid at 26.7 mg/g dried cell weight, a 254% increase.
Conclusions:
- This study reports the first successful gene transfer system for Nitratireductor sp.
- The thioesterase (te) gene was confirmed as responsible for butenoic acid production in Nitratireductor sp. OM-1.
- This work enables further metabolic engineering of Nitratireductor sp. for enhanced butenoic acid synthesis.
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