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Article

Screening of Bacteriophage Encoded Toxic Proteins with a Next Generation Sequencing-Based Assay

1
Department of Bacteriology and Immunology, Medicum, Human Microbiome Research Program, Faculty of Medicine, University of Helsinki, 00290 Helsinki, Finland
2
Division of Clinical Microbiology, HUSLAB, University of Helsinki and Helsinki University Hospital, 00290 Helsinki, Finland
*
Author to whom correspondence should be addressed.
Present address: Department of Chemical Biology & Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, 3584 CG Utrecht, The Netherlands.
Those authors contributed equally to this paper.
Academic Editors: Joana Azeredo and Sílvio Santos
Viruses 2021, 13(5), 750; https://doi.org/10.3390/v13050750
Received: 31 March 2021 / Revised: 17 April 2021 / Accepted: 21 April 2021 / Published: 24 April 2021
Bacteriophage vB_EcoM_fHy-Eco03 (fHy-Eco03 for short) was isolated from a sewage sample based on its ability to infect an Escherichia coli clinical blood culture isolate. Altogether, 32 genes encoding hypothetical proteins of unknown function (HPUFs) were identified from the genomic sequence of fHy-Eco03. The HPUFs were screened for toxic properties (toxHPUFs) with a novel, Next Generation Sequencing (NGS)-based approach. This approach identifies toxHPUF-encoding genes through comparison of gene-specific read coverages in DNA from pooled ligation mixtures before electroporation and pooled transformants after electroporation. The performance and reliability of the NGS screening assay was compared with a plating efficiency-based method, and both methods identified the fHy-Eco03 gene g05 product as toxic. While the outcomes of the two screenings were highly similar, the NGS screening assay outperformed the plating efficiency assay in both reliability and efficiency. The NGS screening assay can be used as a high throughput method in the search for new phage-inspired antimicrobial molecules. View Full-Text
Keywords: bacteriophage; hypothetical proteins of unknown function; next-generation sequencing; toxic protein screen bacteriophage; hypothetical proteins of unknown function; next-generation sequencing; toxic protein screen
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MDPI and ACS Style

Kasurinen, J.; Spruit, C.M.; Wicklund, A.; Pajunen, M.I.; Skurnik, M. Screening of Bacteriophage Encoded Toxic Proteins with a Next Generation Sequencing-Based Assay. Viruses 2021, 13, 750. https://doi.org/10.3390/v13050750

AMA Style

Kasurinen J, Spruit CM, Wicklund A, Pajunen MI, Skurnik M. Screening of Bacteriophage Encoded Toxic Proteins with a Next Generation Sequencing-Based Assay. Viruses. 2021; 13(5):750. https://doi.org/10.3390/v13050750

Chicago/Turabian Style

Kasurinen, Jutta, Cindy M. Spruit, Anu Wicklund, Maria I. Pajunen, and Mikael Skurnik. 2021. "Screening of Bacteriophage Encoded Toxic Proteins with a Next Generation Sequencing-Based Assay" Viruses 13, no. 5: 750. https://doi.org/10.3390/v13050750

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