The enhancement of M13 phage titration by optimizing the origin of replication
| Dublin Core | PKP Metadata Items | Metadata for this Document | |
| 1. | Title | Title of document | The enhancement of M13 phage titration by optimizing the origin of replication |
| 2. | Creator | Author's name, affiliation, country | Mohammad Hossein Darvishali; Department of Genetics and Molecular Biology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran.; Iran, Islamic Republic of |
| 2. | Creator | Author's name, affiliation, country | Mahmood Fadaie; Department of Genetics and Molecular Biology, School of Medicine, Isfahan University of Medical Sciences,Isfahan, Iran.; Iran, Islamic Republic of |
| 2. | Creator | Author's name, affiliation, country | Hossein Khanahmad; Department of Genetics and Molecular Biology, School of Medicine, Isfahan University of Medical Sciences,Isfahan, Iran.; Iran, Islamic Republic of |
| 3. | Subject | Discipline(s) | |
| 3. | Subject | Keyword(s) | Bacteriophage; M13 Phage; Phage titer; Replication origin. |
| 4. | Description | Abstract | Background and purpose: M13KO7, a modified M13 phage variant, carries the p15A replication origin and Tn903 kanamycin resistance gene. This study aimed to optimize M13KO7's replication by substituting the p15A origin with the higher-copy pMB1 origin (500-700 copy numbers). Experimental approach: A 6431-nucleotide fragment from the M13KO7 plasmid lacking the p15A replication origin and kanamycin resistance gene was amplified using a long polymerase chain reaction (PCR). The modified M13AMB1 plasmid was created by adding adenine to the 3’ ends of this fragment and ligating it to the pMB1-containing fragment using T/A cloning. Afterward, to prepare the phage, pM13AMB1 was transformed into E. coli TG1 bacteria, and then, using the PEG-NaCl precipitation, the modified phage was propagated. The modified phage titer was determined utilizing the serial dilution and the qPCR methods, compared with the M13KO7 phage. Findings/Results: The results showed that in the serial dilution method, the titers of modified phage and M13KO7 phage were 4.8 × 1014 and 7 × 1012 pfu/mL, respectively. Besides, the phage titer calculated by the qPCR method for the modified phage was equal to 1.3 × 109 pfu/mL, whereas it was 4.08 × 108 pfu/mL for the M13KO7 phage. Conclusion and implications: This study provides evidence that replication origin replacement led to a significant increase in phage titers. It highlights the importance of replication optimization for molecular biology applications. |
| 5. | Publisher | Organizing agency, location | Isfahan University of Medical Sciences |
| 6. | Contributor | Sponsor(s) | |
| 7. | Date | (YYYY-MM-DD) | 2024-07-07 |
| 8. | Type | Status & genre | Vol 7, No 5 (2012): Supplement (Abstracts of contributed pap |
| 8. | Type | Type | |
| 9. | Format | File format | |
| 10. | Identifier | Uniform Resource Identifier | http://rps.mui.ac.ir/index.php/jrps/article/view/2254 |
| 11. | Source | Title; vol., no. (year) | Research in Pharmaceutical Sciences; Vol 19, No 3 (2024) |
| 12. | Language | English=en | |
| 14. | Coverage | Geo-spatial location, chronological period, research sample (gender, age, etc.) | |
| 15. | Rights | Copyright and permissions |
Copyright (c) 2024 Mohammad Hossein Darvishali, Mahmood Fadaie, Hossein Khanahmad |