A robust strategy for overexpression of DNA polymerase from Thermus aquaticus using an IPTG-independent autoinduction system in a benchtop bioreactor.
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| Format: | Artículo científico |
| Langue: | en |
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Scientific reports
2025
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| author | Laksmi, Fina Amreta Lischer, Kenny Nugraha, Yudhi Violando, Wiga Alif Helbert Nuryana, Isa Khasna, Firyal Nida Nur, Naswandi Ramadhan, Kharisma Panji Tobing, Destrianti Adelina Lumban Hariyatun Hidayat, Iman |
| author_facet | Laksmi, Fina Amreta Lischer, Kenny Nugraha, Yudhi Violando, Wiga Alif Helbert Nuryana, Isa Khasna, Firyal Nida Nur, Naswandi Ramadhan, Kharisma Panji Tobing, Destrianti Adelina Lumban Hariyatun Hidayat, Iman Laksmi, Fina Amreta Lischer, Kenny Nugraha, Yudhi Violando, Wiga Alif Helbert Nuryana, Isa Khasna, Firyal Nida Nur, Naswandi Ramadhan, Kharisma Panji Tobing, Destrianti Adelina Lumban Hariyatun Hidayat, Iman |
| collection | PubMed - marine biology |
| contents | A robust strategy for overexpression of DNA polymerase from Thermus aquaticus using an IPTG-independent autoinduction system in a benchtop bioreactor. Laksmi, Fina Amreta Lischer, Kenny Nugraha, Yudhi Violando, Wiga Alif Helbert Nuryana, Isa Khasna, Firyal Nida Nur, Naswandi Ramadhan, Kharisma Panji Tobing, Destrianti Adelina Lumban Hariyatun Hidayat, Iman Bioreactors Thermus Recombinant Proteins Fermentation Taq Polymerase Isopropyl Thiogalactoside Culture Media Escherichia coli DNA-Directed DNA Polymerase The DNA polymerase derived from Thermus aquaticus is the most widely utilized among various DNA polymerases, indicating its significant economic importance. Consequently, efforts to achieve a substantial yield of Taq DNA polymerase (Taq-pol) are ongoing. The expression of recombinant protein using T7-induced promoters presents challenges in cost-effectiveness, primarily due to the reliance on traditional induction method. Our study aims to enhance cost-efficiency, and scalability of our method for overproducing Taq-pol, particularly in comparison to traditional IPTG-induced techniques, which remain underreported in the current literature. To achieve those purposes, this work integrated the use of (1) a high copy number vector; (2) an optimized chemically defined medium; and (3) optimized fermentation conditions in a 5 L bioreactor. A total of 83.5 mg/L of pure Taq-pol was successfully synthesized in its active form, leading to a 9.7-fold enhancement in protein yield. This was achieved by incorporating glucose, glycerol, and lactose into a defined medium at concentrations of 0.1, 0.6, and 1%, respectively, under specific production conditions in a 5 L bioreactor: 300 rpm, 2 vvm, and 10% inoculant. The data collectively suggest that the strategy serves as a significant foundation for the future advancement of large-scale production of Taq-pol. |
| format | Artículo científico |
| id | pubmed_39966433 |
| institution | PubMed |
| language | en |
| publishDate | 2025 |
| publisher | Scientific reports |
| record_format | pubmed |
| spellingShingle | A robust strategy for overexpression of DNA polymerase from Thermus aquaticus using an IPTG-independent autoinduction system in a benchtop bioreactor. Laksmi, Fina Amreta Lischer, Kenny Nugraha, Yudhi Violando, Wiga Alif Helbert Nuryana, Isa Khasna, Firyal Nida Nur, Naswandi Ramadhan, Kharisma Panji Tobing, Destrianti Adelina Lumban Hariyatun Hidayat, Iman Bioreactors Thermus Recombinant Proteins Fermentation Taq Polymerase Isopropyl Thiogalactoside Culture Media Escherichia coli DNA-Directed DNA Polymerase A robust strategy for overexpression of DNA polymerase from Thermus aquaticus using an IPTG-independent autoinduction system in a benchtop bioreactor. Laksmi, Fina Amreta Lischer, Kenny Nugraha, Yudhi Violando, Wiga Alif Helbert Nuryana, Isa Khasna, Firyal Nida Nur, Naswandi Ramadhan, Kharisma Panji Tobing, Destrianti Adelina Lumban Hariyatun Hidayat, Iman Bioreactors Thermus Recombinant Proteins Fermentation Taq Polymerase Isopropyl Thiogalactoside Culture Media Escherichia coli DNA-Directed DNA Polymerase The DNA polymerase derived from Thermus aquaticus is the most widely utilized among various DNA polymerases, indicating its significant economic importance. Consequently, efforts to achieve a substantial yield of Taq DNA polymerase (Taq-pol) are ongoing. The expression of recombinant protein using T7-induced promoters presents challenges in cost-effectiveness, primarily due to the reliance on traditional induction method. Our study aims to enhance cost-efficiency, and scalability of our method for overproducing Taq-pol, particularly in comparison to traditional IPTG-induced techniques, which remain underreported in the current literature. To achieve those purposes, this work integrated the use of (1) a high copy number vector; (2) an optimized chemically defined medium; and (3) optimized fermentation conditions in a 5 L bioreactor. A total of 83.5 mg/L of pure Taq-pol was successfully synthesized in its active form, leading to a 9.7-fold enhancement in protein yield. This was achieved by incorporating glucose, glycerol, and lactose into a defined medium at concentrations of 0.1, 0.6, and 1%, respectively, under specific production conditions in a 5 L bioreactor: 300 rpm, 2 vvm, and 10% inoculant. The data collectively suggest that the strategy serves as a significant foundation for the future advancement of large-scale production of Taq-pol. |
| title | A robust strategy for overexpression of DNA polymerase from Thermus aquaticus using an IPTG-independent autoinduction system in a benchtop bioreactor. |
| topic | Bioreactors Thermus Recombinant Proteins Fermentation Taq Polymerase Isopropyl Thiogalactoside Culture Media Escherichia coli DNA-Directed DNA Polymerase |
| url | https://pubmed.ncbi.nlm.nih.gov/39966433/ |