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

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Kasurinen , J , Spruit , C M , Wicklund , A , Pajunen , M I & Skurnik , M 2021 , ' Screening of Bacteriophage Encoded Toxic Proteins with a Next Generation Sequencing-Based Assay ' , Viruses (Basel) , vol. 13 , no. 5 , 750 . https://doi.org/10.3390/v13050750

Title: Screening of Bacteriophage Encoded Toxic Proteins with a Next Generation Sequencing-Based Assay
Author: Kasurinen, Jutta; Spruit, Cindy M.; Wicklund, Anu; Pajunen, Maria I.; Skurnik, Mikael
Contributor organization: HUMI - Human Microbiome Research
Department of Bacteriology and Immunology
Medicum
Faculty of Medicine
University of Helsinki
Research Programs Unit
HUSLAB
Glycoscience Group
Biosciences
Helsinki One Health (HOH)
Mikael Skurnik / Principal Investigator
Helsinki University Hospital Area
Date: 2021-05
Language: eng
Number of pages: 16
Belongs to series: Viruses (Basel)
ISSN: 1999-4915
DOI: https://doi.org/10.3390/v13050750
URI: http://hdl.handle.net/10138/332770
Abstract: 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.
Subject: bacteriophage
hypothetical proteins of unknown function
next-generation sequencing
toxic protein screen
ESCHERICHIA-COLI
PHAGE
SEARCH
RECG
IDENTIFICATION
COMPLEXES
DISCOVERY
HELICASE
GENOMES
3111 Biomedicine
11832 Microbiology and virology
Peer reviewed: Yes
Rights: cc_by
Usage restriction: openAccess
Self-archived version: publishedVersion


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