---
title: "Using continuous directed evolution with base editors to map combinatorial TEM-1 antibiotic resist"
id: "pubmed-42763377"
canonical_url: "https://medichelpline.com/clinical-feed/pubmed-42763377"
content_type: "clinical_feed_article"
specialty: "Infectious Disease"
source_name: "PubMed / NCBI"
source_url: "https://pubmed.ncbi.nlm.nih.gov/42763377/"
doi: "10.1007/s10529-026-03791-9"
published_at: "2026-09-20T00:00:00.000Z"
evidence_level: "Journal Article"
license: "CC-BY-NC-4.0 / Informational Use"
---
# Using continuous directed evolution with base editors to map combinatorial TEM-1 antibiotic resist
## Provenance & Clinical Metadata
- **Canonical URL:** https://medichelpline.com/clinical-feed/pubmed-42763377
- **Specialty:** [Infectious Disease](https://medichelpline.com/clinical-feed/infectious-disease.md)
- **Primary Source:** PubMed / NCBI
- **Source URL:** [Original Journal Publication](https://pubmed.ncbi.nlm.nih.gov/42763377/)
- **DOI:** [10.1007/s10529-026-03791-9](https://doi.org/10.1007%2Fs10529-026-03791-9)
- **Published At:** 2026-09-20T00:00:00.000Z
- **Evidence Rating:** Journal Article
## Executive GIST (TL;DR)
- Antimicrobial resistance (AMR) is primarily driven by mutations in antibiotic resistance genes and represents a major threat to human health. - The study adapts **continuous directed evolution (CDE)** combined with **base editor (BE)** gene editing to discover mutations in the TEM-1 β-lactamase gene (**blaTEM-1**). - Researchers performed BE-mediated CDE on TEM-1 carried in a single-copy bacterial artificial chromosome (BAC) vector to identify novel TEM-1 mutants. - Identified mutant genotypes were subsequently validated in a high-copy-number pUC plasmid to confirm sequence and phenotype. - The team investigated combinatorial effects by combining CDE-generated TEM-1 phenotypes with previously reported TEM-1 variants. - A strain of Escherichia coli carrying a vector with 10 TEM-1 mutations exhibited a cefotaxime minimum inhibitory concentration (MIC) of 1280 μg/mL, reported by the authors as the highest value observed to date. - The study highlights the utility of gene editing approaches to illuminate the molecular basis of antibiotic resistance and to identify high-resistance genotypes. - Keywords and methodological focus include **Base editor**, **Continuous directed evolution**, **Minimum inhibitory concentration**, **Recombineering**, and **TEM-1**. - Experimental model systems and concepts referenced include Escherichia coli, bacterial artificial chromosomes, pUC plasmids, beta-lactamases, and cefotaxime susceptibility testing. - Details on experimental protocols, specific mutation identities, evolution parameters, and broader clinical implications were not fully reported in the source abstract and would require consulting the full article for complete methods and data.
## Clinical Analysis & Structured Key Points
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R. China."), [Jiamei Wang](https://pubmed.ncbi.nlm.nih.gov/?term=Wang+J&cauthor_id=42763377)[ 1 ](https://pubmed.ncbi.nlm.nih.gov/42763377/#full-view-affiliation-1 "Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China."), [Di Zhang](https://pubmed.ncbi.nlm.nih.gov/?term=Zhang+D&cauthor_id=42763377)[ 1 ](https://pubmed.ncbi.nlm.nih.gov/42763377/#full-view-affiliation-1 "Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China."), [Guangdong Shang](https://pubmed.ncbi.nlm.nih.gov/?term=Shang+G&cauthor_id=42763377)[ 2 ](https://pubmed.ncbi.nlm.nih.gov/42763377/#full-view-affiliation-2 "Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China. shanggd@hotmail.com.") Affiliations Expand ### Affiliations * 1 Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China. * 2 Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China. shanggd@hotmail.com. * PMID: **42763377** * DOI: [ 10.1007/s10529-026-03791-9 ](https://doi.org/10.1007/s10529-026-03791-9) Item in Clipboard # Coupling continuous directed evolution with base editors identifies combinatorial antibiotic resistance Hong Wang et al. Biotechnol Lett. 2026. Show details Display options Display options Format Abstract PubMed PMID Biotechnol Lett Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Biotechnol+Lett%22%5Bjour%5D&sort=date&sort_order=desc) * [ Search in NLM Catalog ](https://www.ncbi.nlm.nih.gov/nlmcatalog?term=%22Biotechnol+Lett%22%5BTitle+Abbreviation%5D) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) . 2026 Sep 20;48(5):117. doi: 10.1007/s10529-026-03791-9. ### Authors [Hong Wang](https://pubmed.ncbi.nlm.nih.gov/?term=Wang+H&cauthor_id=42763377)[ 1 ](https://pubmed.ncbi.nlm.nih.gov/42763377/#short-view-affiliation-1 "Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China."), [Jiamei Wang](https://pubmed.ncbi.nlm.nih.gov/?term=Wang+J&cauthor_id=42763377)[ 1 ](https://pubmed.ncbi.nlm.nih.gov/42763377/#short-view-affiliation-1 "Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China."), [Di Zhang](https://pubmed.ncbi.nlm.nih.gov/?term=Zhang+D&cauthor_id=42763377)[ 1 ](https://pubmed.ncbi.nlm.nih.gov/42763377/#short-view-affiliation-1 "Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China."), [Guangdong Shang](https://pubmed.ncbi.nlm.nih.gov/?term=Shang+G&cauthor_id=42763377)[ 2 ](https://pubmed.ncbi.nlm.nih.gov/42763377/#short-view-affiliation-2 "Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China. shanggd@hotmail.com.") ### Affiliations * 1 Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China. * 2 Jiangsu Key Laboratory for Pathogens and Ecosystems, College of Life Sciences, Jiangsu Province, Nanjing Normal University, No.1 Wenyuan Rd, Xixia District, Nanjing, 210023, P. R. China. shanggd@hotmail.com. * PMID: **42763377** * DOI: [ 10.1007/s10529-026-03791-9 ](https://doi.org/10.1007/s10529-026-03791-9) Item in Clipboard Cite Display options Display options Format Abstract PubMed PMID ## Abstract Antimicrobial resistance (AMR), mainly caused by the mutation of antibiotic resistance genes, poses a great threat to human health. Clinically, TEM-1 β-lactamase encoding blaTEM-1 gene is the most frequently occurred antibiotic resistance gene. Identification, characterization, and monitor of high antibiotic resistance blaTEM-1 mutants are crucial to AMR research and would provide the guidance for the development of next-generation antibiotics. To this end, we report here the adaption of the continuous directed evolution (CDE) along with base editor (BE) gene editing strategy to discover TEM-1 mutants. Firstly, TEM-1 mutants were identified in single-copy bacterial artificial chromosome vector via BE-mediated CDE. Then each mutant genotype was verified in high copy number pUC plasmid. Finally, combinatorial antibiotic resistance was observed when the TEM-1 phenotypes generated in CDE were combined with that of the reported TEM-1 variants. The Escherichia coli strain harboring the vector cloned with 10 TEM-1 mutations showed a minimum inhibitory concentration of 1280 μg/mL to antibiotic cefotaxime, which is the highest data reported thus far. The research highlights the application of gene editing methods to provide insight into the molecular basis of antibiotic resistance. **Keywords:** Base editor; Continuous directed evolution; Minimum inhibitory concentration; Recombineering; TEM-1. © 2026. The Author(s), under exclusive licence to Springer Nature B.V. [PubMed Disclaimer](https://pubmed.ncbi.nlm.nih.gov/disclaimer/) ## Conflict of interest statement Declarations. Conflict of interest: The authors report no conflicts of interest. ## References 1. 1. Ambler RP, Coulson AF, Frere JM et al (1991) A standard numbering scheme for the class A β-lactamases. Biochem J 276:269–270. - [DOI](https://doi.org/10.1042/bj2760269) - [PubMed](https://pubmed.ncbi.nlm.nih.gov/2039479/) - [PMC](https://pmc.ncbi.nlm.nih.gov/articles/1151176/) 2. 1. Bershtein S, Tawfik DS (2008) Ohno’s model revisited: measuring the frequency of potentially adaptive mutations under various mutational drifts. Mol Biol Evol 25:2311–2318. - [DOI](https://doi.org/10.1093/molbev/msn174) - [PubMed](https://pubmed.ncbi.nlm.nih.gov/18687656/) 3. 1. Chen Z, Ling W, Shang G (2016) Recombineering and I-SceI-mediated Pseudomonas putida KT2440 scarless gene deletion. FEMS Microbiol Lett 363:fnw231. - [DOI](https://doi.org/10.1093/femsle/fnw231) - [PubMed](https://pubmed.ncbi.nlm.nih.gov/27765807/) 4. 1. Dance A (2024) Five ways science is tackling the antibiotic resistance crisis. Nature 632:494–496. - [DOI](https://doi.org/10.1038/d41586-024-02601-4) - [PubMed](https://pubmed.ncbi.nlm.nih.gov/39138306/) 5. 1. Datsenko KA, Wanner BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 97:6640–6645. - [DOI](https://doi.org/10.1073/pnas.120163297) - [PubMed](https://pubmed.ncbi.nlm.nih.gov/10829079/) - [PMC](https://pmc.ncbi.nlm.nih.gov/articles/18686/) Show all 36 references ## MeSH terms * Anti-Bacterial Agents / pharmacology Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Anti-Bacterial+Agents%2Fpharmacology%22%5BMeSH%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Anti-Bacterial+Agents) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Cefotaxime / pharmacology Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Cefotaxime%2Fpharmacology%22%5BMeSH%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Cefotaxime) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Chromosomes, Artificial, Bacterial / genetics Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Chromosomes%2C+Artificial%2C+Bacterial%2Fgenetics%22%5BMeSH%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Chromosomes%2C+Artificial%2C+Bacterial) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Directed Molecular Evolution* / methods Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Directed+Molecular+Evolution%2Fmethods%22%5BMAJR%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Directed+Molecular+Evolution) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Drug Resistance, Bacterial* / genetics Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Drug+Resistance%2C+Bacterial%2Fgenetics%22%5BMAJR%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Drug+Resistance%2C+Bacterial) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Escherichia coli* / drug effects Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Escherichia+coli%2Fdrug+effects%22%5BMAJR%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Escherichia+coli) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Escherichia coli* / genetics Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Escherichia+coli%2Fgenetics%22%5BMAJR%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Escherichia+coli) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Gene Editing* / methods Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Gene+Editing%2Fmethods%22%5BMAJR%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Gene+Editing) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Microbial Sensitivity Tests Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Microbial+Sensitivity+Tests%22%5BMeSH%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Microbial+Sensitivity+Tests) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * Mutation Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22Mutation%22%5BMeSH%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=Mutation) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) * beta-Lactamases* / genetics Actions * [ Search in PubMed ](https://pubmed.ncbi.nlm.nih.gov/?term=%22beta-Lactamases%2Fgenetics%22%5BMAJR%5D&sort=date&sort_order=desc) * [ Search in MeSH ](https://www.ncbi.nlm.nih.gov/mesh?term=beta-Lactamases) * [ Add to Search ](https://pubmed.ncbi.nlm.nih.gov/42763377/) ## 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