# Harnessing Broad-Specificity Phage Recombinases for Universal Bacterial Editing platforms

> **NIH NIH DP2** · UNIVERSITY OF ROCHESTER · 2024 · $453,900

## Abstract

PROJECT SUMMARY
The vast array of genetic elements within a bacterial genome dictates its potential to cause disease. These
elements influence virulence, antibiotic susceptibility, and the ability to evade the immune response.
Understanding and manipulating these genetic components are crucial for identifying the drivers of bacterial
pathogenicity. However, a significant challenge arises because efficient genetic systems are typically limited to
laboratory-adapted strains. This limitation leaves many virulent clinical pathogens genetically intractable. To
address this challenge, we developed a genome editing platform on phage recombinase that works across a
broad range of species. Editing bacteria with phage recombinases, a process termed recombineering, has been
transformative in E coli. However, its reliance on host factors causes species-specificity that has limited its
broader impact. We discovered a phage protein that is conserved across bacterial phyla that can used to identify
broadly functioning phage recombinase systems. Leveraging this protein, we developed a recombineering
system that operates in both Gram-positive and Gram-negative bacteria. To further explore the potential of
phage-based genetic engineering, we have established a high-throughput selection to identify phage
recombinases from genomic material without relying on homology to existing recombinases. Coupling the
ubiquity of recombination modules in dsDNA phages with the vast diversity of genetic information encoded by
phage, we anticipate identifying numerous new phage-based recombination systems. Initial results have
demonstrated levels of genetic editing in the important human pathogen Staphylococcus aureus comparable to
recombineering in E coli. This work will transform the way we genetically manipulate bacteria, enabling us to
directly probe the genetic elements driving pathogenicity in clinically relevant strains.

## Key facts

- **NIH application ID:** 10949315
- **Project number:** 1DP2AI184732-01
- **Recipient organization:** UNIVERSITY OF ROCHESTER
- **Principal Investigator:** Andrew Varble
- **Activity code:** DP2 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $453,900
- **Award type:** 1
- **Project period:** 2024-08-13 → 2029-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10949315

## Citation

> US National Institutes of Health, RePORTER application 10949315, Harnessing Broad-Specificity Phage Recombinases for Universal Bacterial Editing platforms (1DP2AI184732-01). Retrieved via AI Analytics 2026-06-12 from https://api.ai-analytics.org/grant/nih/10949315. Licensed CC0.

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