Awakening prime editing for precision engineering of probiotic Escherichia coli Nissle 1917 DOI Creative Commons

Peiru Chen,

Ying Wei, Xin Li

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 28, 2024

Abstract CRISPR-Cas systems are transforming precision medicine with engineered probiotics as next-generation diagnostics and therapeutics. To promote human health treat disease, engineering probiotic bacteria demands maximal versatility to enable non-natural functionalities while minimizing undesired genomic interferences. Here, we present a streamlined prime editing approach tailored for Escherichia coli Nissle 1917 utilizing only essential genetic modules an optimized workflow. This was realized by assembling editor consisting of the system from Streptococcus pyogenes its native codons codon-optimized reverse transcriptase, orchestrating induction levels. As result, achieved all types in every individual round experiments efficiencies 25.0%, 52.0% 66.7% DNA deletion, insertion, substitution, respectively. A comprehensive evaluation off-target effects revealed significant reduction unintended mutations, particularly comparison two different base methods. Leveraging system, developed barcoding strain tracking antibiotic-resistance-gene-free platform functionalities. Our strategy awakens back-to-basics devoid complex or extraneous designs, paving way future innovations probiotics.

Language: Английский

Biomanufacturing of value-added chemicals from lignin DOI
Arren Liu, Dylan Ellis, Apurv Mhatre

et al.

Current Opinion in Biotechnology, Journal Year: 2024, Volume and Issue: 89, P. 103178 - 103178

Published: Aug. 3, 2024

Language: Английский

Citations

4
Precision engineering of the probiotic Escherichia coli Nissle 1917 with prime editing DOI Creative Commons

P. Chen,

Ying Wei, Xin Li

et al.

Applied and Environmental Microbiology, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 31, 2025

ABSTRACT CRISPR-Cas systems are transforming precision medicine with engineered probiotics as next-generation diagnostics and therapeutics. To promote human health treat disease, engineering probiotic bacteria demands maximal versatility to enable non-natural functionalities while minimizing undesired genomic interferences. Here, we present a streamlined prime editing approach tailored for Escherichia coli Nissle 1917 utilizing only essential genetic modules, including Cas9 nickase from Streptococcus pyogenes , codon-optimized reverse transcriptase, guide RNA, an optimized workflow longer induction. As result, achieved all types of in every individual round experiments efficiencies 25.0%, 52.0%, 66.7% DNA deletion, insertion, substitution, respectively. A comprehensive evaluation off-target effects revealed significant reduction unintended mutations, particularly comparison two different base methods. Leveraging the system, inserted unique sequence barcode edited strain established antibiotic-resistance-gene-free platform functionalities. Our strategy presents system that can be readily implemented any laboratories basic CRISPR setups, paving way future innovations probiotics. IMPORTANCE One ultimate goal gene is introduce designed variations at specific loci living organisms minimal interferences genome. Achieving this especially critical creating therapeutics diseases. In endeavor, report customized 1917. With such developed barcoding tracking strains, built We provide not powerful but also new insights into advancement innovative systems.

Language: Английский

Citations

0
Awakening prime editing for precision engineering of probiotic Escherichia coli Nissle 1917 DOI Creative Commons

Peiru Chen,

Ying Wei, Xin Li

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 28, 2024

Abstract CRISPR-Cas systems are transforming precision medicine with engineered probiotics as next-generation diagnostics and therapeutics. To promote human health treat disease, engineering probiotic bacteria demands maximal versatility to enable non-natural functionalities while minimizing undesired genomic interferences. Here, we present a streamlined prime editing approach tailored for Escherichia coli Nissle 1917 utilizing only essential genetic modules an optimized workflow. This was realized by assembling editor consisting of the system from Streptococcus pyogenes its native codons codon-optimized reverse transcriptase, orchestrating induction levels. As result, achieved all types in every individual round experiments efficiencies 25.0%, 52.0% 66.7% DNA deletion, insertion, substitution, respectively. A comprehensive evaluation off-target effects revealed significant reduction unintended mutations, particularly comparison two different base methods. Leveraging system, developed barcoding strain tracking antibiotic-resistance-gene-free platform functionalities. Our strategy awakens back-to-basics devoid complex or extraneous designs, paving way future innovations probiotics.

Language: Английский

Citations

0