ArticleMolecular systems biology2020
CRISPR/Cas9 recombineering-mediated deep mutational scanning of essential genes in Escherichia coli.
Article in Molecular systems biology, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 27 papers.
What it found
Each row is one number read from the abstract, on the scale the paper reported it, with its interval. Left of the dashed line favours the treatment, right favours the comparator. Under each row is the sentence it came from. New to these charts? A ten-minute tutorial.
The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
Neither the registry nor the abstract names a trial number. If this is a trial report, that itself is worth knowing.
Who cites it
27 citing papers in PubMed, 47 citations in OpenAlex.
- Functional profiling of 2,193 ASS1 missense variants: Insights into variant pathogenicity and epistatic interactions in citrullinemia type I.PLoS genetics · 2026Article
- Enhancement of single-stranded template annealing activity by Rad52 during repair of CRISPR-induced dsDNA breaks.Nucleic acids research · 2026Article
- D-LIM: A neural network for interpretable gene-gene interactions.PLoS computational biology · 2026Article
- Multilayered regulation by RNA thermometers enables precise control of Cas9 expression in E. coli.Nucleic acids research · 2026Article
- High-throughput conjugation reveals strain specific recombination patterns enabling precise trait mapping in Escherichia coli.PLoS genetics · 2025Article
- Bacterial Systematic Genetics and Integrated Multi-Omics: Beyond Static Genomics Toward Predictive Models.International journal of molecular sciences · 2025Review
- EASY-edit: a toolbox for high-throughput single-step custom genetic editing in bacteria.Nucleic acids research · 2025Article
- Idiosyncratic evolvability among single-point ribosomal mutants towards multi-aminoglycoside resistance.PLoS genetics · 2025Article
- Enhancing glucaric acid production fromApplied and environmental microbiology · 2024Article
- ORBIT for E. coli: kilobase-scale oligonucleotide recombineering at high throughput and high efficiency.Nucleic acids research · 2024Article
- CRISPR-Cas9-assisted genome editing inbioRxiv : the preprint server for biology · 2024Article
- Recent advances in genome-scale engineering inEngineering microbiology · 2024Review
- Deep mutational scanning reveals the molecular determinants of RNA polymerase-mediated adaptation and tradeoffs.Nature communications · 2023Article
- ShinyBioHEAT: an interactive shiny app to identify phenotype driver genes in E.coli and B.subtilis.Bioinformatics (Oxford, England) · 2023Article
- Cas9 off-target binding to the promoter of bacterial genes leads to silencing and toxicity.Nucleic acids research · 2023Article
- Article
- Mutational fitness landscape and drug resistance.Current opinion in structural biology · 2023Review
- Deep mutational scanning: A versatile tool in systematically mapping genotypes to phenotypes.Frontiers in genetics · 2023Review
- CRISPR-based oligo recombineering prioritizes apicomplexan cysteines for drug discovery.Nature microbiology · 2022Article
- Interpreting protein variant effects with computational predictors and deep mutational scanning.Disease models & mechanisms · 2022Review
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
6 authors at 2 institutions in 3 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Deep mutational scanning can provide significant insights into the function of essential genes in bacteria. Here, we developed a high-throughput method for mutating essential genes of Escherichia coli in their native genetic context. We used Cas9-mediated recombineering to introduce a library of mutations, created by error-prone PCR, within a gene fragment on the genome using a single gRNA pre-validated for high efficiency. Tracking mutation frequency through deep sequencing revealed biases in the position and the number of the introduced mutations. We overcame these biases by increasing the homology arm length and blocking mismatch repair to achieve a mutation efficiency of 85% for non-essential genes and 55% for essential genes. These experiments also improved our understanding of poorly characterized recombineering process using dsDNA donors with single nucleotide changes. Finally, we applied our technology to target rpoB, the beta subunit of RNA polymerase, to study resistance against rifampicin. In a single experiment, we validate multiple biochemical and clinical observations made in the previous decades and provide insights into resistance compensation with the study of double mutants.
Indexed as
Identifiers
What OpenQuestion holds
Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.