Continuous Secretion of Human Epidermal Growth Factor Based on Escherichia coli Biofilm
Abstract
:1. Introduction
2. Materials and Methods
2.1. Strains and Plasmids
2.2. Media and Growth Conditions
2.3. Fermentation and Analytical Methods
2.4. Characterization of Biofilm Formation
2.5. Congo Red Assay
2.6. Confocal Laser Scanning Microscope
3. Results
3.1. Screening Results of High Production Strain
3.2. Effect of Gene Modification on Biofilm Formation
3.3. Effects of Overexpression of Biofilm-Related Genes on hEGF Secretion
3.4. Production of hEGF by Biofilm-Immobilized Continuous Fermentation
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Strains or Plasmids | Relevant Characteristics | Sources | |
---|---|---|---|
Strains | E. coli BL21(DE3) | Host strain | Invitrogen |
BL21-hEGF-C1 | Genome-integrated expression of hEGF in one locus | This study | |
BL21-hEGF-C2 | Genome-integrated expression of hEGF in two loci | This study | |
BL21-hEGF-C3 | Genome-integrated expression of hEGF in three loci | This study | |
BL21-hEGF-C4 | Genome-integrated expression of hEGF in four loci | This study | |
BL21-hEGF-C5 | Genome-integrated expression of hEGF in five loci | This study | |
BL21-hEGF-dgcC | Plasmid-based expression of dgcC | This study | |
BL21-hEGF-csgD | Plasmid-based expression of csgD | This study | |
BL21-hEGF-bcsA | Plasmid-based expression of bcsA | This study | |
BL21-hEGF-bcsB | Plasmid-based expression of bcsB | This study | |
BL21-hEGF-pBb-bcsB * | Plasmid-based expression of bcsB | This study | |
Plasmids | pET30a-PelB-hEGF | hEGF expression | [15] |
pCas | CRISPR editing | [16] | |
pTarget | CRISPR editing | [16] | |
pET28a | Expression vector | Invitrogen | |
pBbE1a | Expression vector | [15] | |
pET28a-dgcC | Plasmid-based expression of dgcC | This study | |
pET28a-csgD | Plasmid-based expression of csgD | This study | |
pET28a-bcsA | Plasmid-based expression of bcsA | This study | |
pET28a-bcsB | Plasmid-based expression of bcsB | This study | |
pBbE1a-bcsB | Plasmid-based expression of bcsB | [15] |
Strains | Congo Red Binding Ratio |
---|---|
BL21-hEGF-C5 | 0.37 |
BL21-hEGF-dgcC | 0.51 |
BL21-hEGF-csgD | 0.47 |
BL21-hEGF-bcsA | 0.69 |
BL21-hEGF-bcsB | 0.72 |
BL21-hEGF-pBb-bcsB | 0.51 |
Strains | hEGF Secretion of the Modified Strains Compared to the Control Strain (%) | |||||||
---|---|---|---|---|---|---|---|---|
1st | 2nd | 3rd | 4th | 5th | 6th | 7th | Average | |
BL21-hEGF-dgcC | −3.5 | 8.4 | 10.2 | 8.9 | 7.7 | 10.3 | 8.7 | 7.2 |
BL21-hEGF-csgD | 8.8 | 20.3 | 33.9 | 27.0 | 30.4 | 21.8 | 24.1 | 23.8 |
BL21-hEGF-bcsA | 3.8 | 24.5 | 18.1 | 13.9 | 14.5 | 15.3 | 11.7 | 14.5 |
BL21-hEGF-bcsB | 16.5 | 24.5 | 36.7 | 32.7 | 32.2 | 29.6 | 24.1 | 28.0 |
BL21-hEGF-pBb- bcsB | 11.5 | 19.2 | 26.4 | 25.3 | 20.4 | 21.0 | 20.4 | 20.6 |
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Zhang, C.; Liao, J.; Li, Y.; Liu, S.; Li, M.; Zhang, D.; Wang, Z.; Liu, D.; Ying, H. Continuous Secretion of Human Epidermal Growth Factor Based on Escherichia coli Biofilm. Fermentation 2024, 10, 202. https://doi.org/10.3390/fermentation10040202
Zhang C, Liao J, Li Y, Liu S, Li M, Zhang D, Wang Z, Liu D, Ying H. Continuous Secretion of Human Epidermal Growth Factor Based on Escherichia coli Biofilm. Fermentation. 2024; 10(4):202. https://doi.org/10.3390/fermentation10040202
Chicago/Turabian StyleZhang, Chong, Jinglin Liao, Yuancong Li, Shuli Liu, Mengting Li, Di Zhang, Zhenyu Wang, Dong Liu, and Hanjie Ying. 2024. "Continuous Secretion of Human Epidermal Growth Factor Based on Escherichia coli Biofilm" Fermentation 10, no. 4: 202. https://doi.org/10.3390/fermentation10040202