ArticleACS synthetic biology2020
Saturation Mutagenesis Genome Engineering of Infective ΦX174 Bacteriophage
Article in ACS synthetic biology, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 11 papers.
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Who cites it
11 citing papers in PubMed.
- Synthetic Biology-Based Engineering Living Therapeutics for Antimicrobial Application.Exploration (Beijing, China) · 2025Article
- Engineering bacteriophages through deep mining of metagenomic motifs.Science advances · 2025Article
- Applications of bacteriophages in precision engineering of the human gut microbiome.Engineering microbiology · 2025Review
- Engineering Phages to Fight Multidrug-Resistant Bacteria.Chemical reviews · 2025Review
- Cell-free synthesis of infective phages fromSynthetic biology (Oxford, England) · 2024Article
- High-throughput approaches to understand and engineer bacteriophages.Trends in biochemical sciences · 2023Review
- LibGENiE - A bioinformatic pipeline for the design of information-enriched enzyme libraries.Computational and structural biotechnology journal · 2023Article
- Oligo Pools as an Affordable Source of Synthetic DNA for Cost-Effective Library Construction in Protein- and Metabolic Pathway Engineering.Chembiochem : a European journal of chemical biology · 2022Review
- Article
- ΦX174 Attenuation by Whole-Genome Codon Deoptimization.Genome biology and evolution · 2021Article
- A Method for User-defined Mutagenesis by Integrating Oligo Pool Synthesis Technology with Nicking Mutagenesis.Bio-protocol · 2020Article
Corrections and comments
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Authors and funding
8 authors.
Funding
Abstract
Here we present a novel protocol for the construction of saturation single-site-and massive multisite-mutant libraries of a bacteriophage. We segmented the ΦX174 genome into 14 nontoxic and nonreplicative fragments compatible with Golden Gate assembly. We next used nicking mutagenesis with oligonucleotides prepared from unamplified oligo pools with individual segments as templates to prepare near-comprehensive single-site mutagenesis libraries of genes encoding the F capsid protein (421 amino acids scanned) and G spike protein (172 amino acids scanned). Libraries possessed greater than 99% of all 11 860 programmed mutations. Golden Gate cloning was then used to assemble the complete ΦX174 mutant genome and generate libraries of infective viruses. This protocol will enable reverse genetics experiments for studying viral evolution and, with some modifications, can be applied for engineering therapeutically relevant bacteriophages with larger genomes.
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Registered trials
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