Development of a Molasses-Based Medium for <i>Agrobacterium tumefaciens</i> Fermentation for Application in Plant-Based Recombinant Protein Production
The <i>Agrobacterium</i>-mediated transient gene expression system is a rapid and efficient method for heterologous recombinant protein expression in plants. The fermentation of genetically modified <i>Agrobacterium tumefaciens</i> is an important step in increasing the effic...
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| Format: | Article |
| Language: | English |
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MDPI AG
2023-02-01
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| Series: | Fermentation |
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| Online Access: | https://www.mdpi.com/2311-5637/9/2/149 |
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| author | Nisit Watthanasakphuban Luan Van Nguyen Yu-Shen Cheng Pau-Loke Show Malinee Sriariyanun Mattheos Koffas Kittipong Rattanaporn |
| author_facet | Nisit Watthanasakphuban Luan Van Nguyen Yu-Shen Cheng Pau-Loke Show Malinee Sriariyanun Mattheos Koffas Kittipong Rattanaporn |
| author_sort | Nisit Watthanasakphuban |
| collection | DOAJ |
| description | The <i>Agrobacterium</i>-mediated transient gene expression system is a rapid and efficient method for heterologous recombinant protein expression in plants. The fermentation of genetically modified <i>Agrobacterium tumefaciens</i> is an important step in increasing the efficiency of recombinant protein production in plants. However, the limitation of this system that makes it economically non-competitive for industrial-scale applications is the <i>Agrobacterium</i> suspension production cost. In this study, the utilization of sugarcane molasses as an alternative low-cost source of carbon at a concentration of 8.7 g/L and nitrogen at a concentration of 2.4 g/L for <i>Agrobacterium</i> cultivation was investigated. Molasses pretreatment using sulfuric acid (SA) was applied before fermentation, and it resulted in a maximum specific growth rate of 0.232 ± 0.0063 h<sup>−1</sup> in the <i>A. tumefaciens</i> EHA105 culture. The supplementation of antibiotics in the molasses-based medium was shown to be unnecessary for plasmid maintenance during fermentation in both <i>Agrobacterium</i> strains, which helped to reduce the production cost. We evaluated recombinant protein production using an <i>Agrobacterium</i> culture without antibiotic supplementation in the growth media by demonstrating green fluorescent protein expression in wild-type <i>Nicotiana benthamiana</i> leaves. In the evaluation of the culture medium cost, the molasses-based medium cost was 6.1 times lower than that of LB. Finally, this study demonstrated that the newly developed molasses-based medium for <i>Agrobacterium</i> fermentation is a feasible and effective medium for transient recombinant protein production in plant tissues. |
| first_indexed | 2024-03-11T08:50:43Z |
| format | Article |
| id | doaj.art-4824b2e614e44633894d55fb0e9cb41d |
| institution | Directory Open Access Journal |
| issn | 2311-5637 |
| language | English |
| last_indexed | 2024-03-11T08:50:43Z |
| publishDate | 2023-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Fermentation |
| spelling | doaj.art-4824b2e614e44633894d55fb0e9cb41d2023-11-16T20:26:08ZengMDPI AGFermentation2311-56372023-02-019214910.3390/fermentation9020149Development of a Molasses-Based Medium for <i>Agrobacterium tumefaciens</i> Fermentation for Application in Plant-Based Recombinant Protein ProductionNisit Watthanasakphuban0Luan Van Nguyen1Yu-Shen Cheng2Pau-Loke Show3Malinee Sriariyanun4Mattheos Koffas5Kittipong Rattanaporn6Fermentation Technology Research Center, Department of Biotechnology, Faculty of Agro-Industry, Kasetsart University, Bangkok 10900, ThailandDepartment of Biotechnology, Faculty of Agro-Industry, Kasetsart University, Bangkok 10900, ThailandDepartment of Chemical and Materials Engineering, National Yunlin University of Science and Technology, Douliu, Yunlin 64002, TaiwanDepartment of Chemical and Environmental Engineering, Faculty of Engineering, University of Nottingham Malaysia, Semenyih 43500, Selangor, MalaysiaBiorefinery and Process Automation Engineering Center, Department of Chemical and Process Engineering, The Sirindhorn International Thai-German Graduate School of Engineering, King Mongkut’s University of Technology North Bangkok, Bangkok 10800, ThailandDepartment of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, NY 12180, USAFermentation Technology Research Center, Department of Biotechnology, Faculty of Agro-Industry, Kasetsart University, Bangkok 10900, ThailandThe <i>Agrobacterium</i>-mediated transient gene expression system is a rapid and efficient method for heterologous recombinant protein expression in plants. The fermentation of genetically modified <i>Agrobacterium tumefaciens</i> is an important step in increasing the efficiency of recombinant protein production in plants. However, the limitation of this system that makes it economically non-competitive for industrial-scale applications is the <i>Agrobacterium</i> suspension production cost. In this study, the utilization of sugarcane molasses as an alternative low-cost source of carbon at a concentration of 8.7 g/L and nitrogen at a concentration of 2.4 g/L for <i>Agrobacterium</i> cultivation was investigated. Molasses pretreatment using sulfuric acid (SA) was applied before fermentation, and it resulted in a maximum specific growth rate of 0.232 ± 0.0063 h<sup>−1</sup> in the <i>A. tumefaciens</i> EHA105 culture. The supplementation of antibiotics in the molasses-based medium was shown to be unnecessary for plasmid maintenance during fermentation in both <i>Agrobacterium</i> strains, which helped to reduce the production cost. We evaluated recombinant protein production using an <i>Agrobacterium</i> culture without antibiotic supplementation in the growth media by demonstrating green fluorescent protein expression in wild-type <i>Nicotiana benthamiana</i> leaves. In the evaluation of the culture medium cost, the molasses-based medium cost was 6.1 times lower than that of LB. Finally, this study demonstrated that the newly developed molasses-based medium for <i>Agrobacterium</i> fermentation is a feasible and effective medium for transient recombinant protein production in plant tissues.https://www.mdpi.com/2311-5637/9/2/149sugarcane molassesmolasses pretreatmentfermentation<i>Agrobacterium tumefaciens</i>transient protein expression |
| spellingShingle | Nisit Watthanasakphuban Luan Van Nguyen Yu-Shen Cheng Pau-Loke Show Malinee Sriariyanun Mattheos Koffas Kittipong Rattanaporn Development of a Molasses-Based Medium for <i>Agrobacterium tumefaciens</i> Fermentation for Application in Plant-Based Recombinant Protein Production Fermentation sugarcane molasses molasses pretreatment fermentation <i>Agrobacterium tumefaciens</i> transient protein expression |
| title | Development of a Molasses-Based Medium for <i>Agrobacterium tumefaciens</i> Fermentation for Application in Plant-Based Recombinant Protein Production |
| title_full | Development of a Molasses-Based Medium for <i>Agrobacterium tumefaciens</i> Fermentation for Application in Plant-Based Recombinant Protein Production |
| title_fullStr | Development of a Molasses-Based Medium for <i>Agrobacterium tumefaciens</i> Fermentation for Application in Plant-Based Recombinant Protein Production |
| title_full_unstemmed | Development of a Molasses-Based Medium for <i>Agrobacterium tumefaciens</i> Fermentation for Application in Plant-Based Recombinant Protein Production |
| title_short | Development of a Molasses-Based Medium for <i>Agrobacterium tumefaciens</i> Fermentation for Application in Plant-Based Recombinant Protein Production |
| title_sort | development of a molasses based medium for i agrobacterium tumefaciens i fermentation for application in plant based recombinant protein production |
| topic | sugarcane molasses molasses pretreatment fermentation <i>Agrobacterium tumefaciens</i> transient protein expression |
| url | https://www.mdpi.com/2311-5637/9/2/149 |
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