# Streamlining temperate phage engineering to facilitate precise in situ manipulation of gut microbiota

> **NIH NIH K99** · NEW YORK UNIVERSITY SCHOOL OF MEDICINE · 2022 · $100,000

## Abstract

PROJECT SUMMARY
This project aims to improve the overall accessibility of precise tools for manipulating mammalian microbiota
within native gut environments, i.e., in situ. Despite the rich microbial diversity that mammalian gut communities
natively acquire and maintain over time, the feasibility of experimentally and therapeutically altering their
composition with antibiotics or fecal transplants has been clearly documented. However, these widely available
treatments are found to broadly alter microbiome composition and this has limited their practical utility for
determining how particular populations of microbiota influence their host’s health. Alternative approaches that
allow for precise elimination or genetic editing of endogenous gut commensals in situ have recently been
demonstrated using horizontally transmissible mobile genetic elements (MGEs), including temperate phages,
but are not yet widely established. Phages are thought to be among the most precise MGEs for in situ microbiome
manipulation given that the majority of studies on their host range have failed to detect cross-genus infectivity.
Studies proposed in this application harness the natural abundance and precision of temperate phages and will
establish generalizable methods to re-engineer them for more reliable use in microbiome editing applications.
Namely, my proposed directed evolution and rational engineering approaches will streamline the generation of
(1) lethal virulent mutants that are obligately lytic and capable of superinfecting their lysogenic kin, as well as (2)
non-lethal temperate phage derivatives that can eliminate endogenous prophages or stably lysogenize at
elevated frequencies. The initial mentored K99 phase of this research includes proof-of-principle engineering
experiments with lambda, the most well-characterized temperate phage of Escherichia coli. In addition to
providing me with hands-on lambda experience, this vital K99 training will prepare me for independent research
activities during the R00 phase and throughout my future career by allowing me to become skilled in key
techniques—such as large-scale DNA assembly in yeast for construction of phage genome libraries and whole
phage genome sequencing—that can be applied to the study of non-model microbiota. Training will be
supervised by two mentors with the requisite resources and experience to ensure my success on this project,
Dr. Jef Boeke and Dr. Marcus Noyes, as well as four advisory committee members with complementary
expertise. Career-oriented guidance from my mentors and advisors, along with career development activities
during the K99 phase that include formal coursework on grant writing and project management, will further
facilitate my transition to the R00 phase and my long-term productivity as an independent academic investigator.
Ultimately, completion of this proposed project will open new avenues for well-controlled experimentation in the
microbiome field while providing fundamental t...

## Key facts

- **NIH application ID:** 10507364
- **Project number:** 1K99GM147604-01
- **Recipient organization:** NEW YORK UNIVERSITY SCHOOL OF MEDICINE
- **Principal Investigator:** Gregory William Goldberg
- **Activity code:** K99 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $100,000
- **Award type:** 1
- **Project period:** 2022-09-07 → 2024-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10507364

## Citation

> US National Institutes of Health, RePORTER application 10507364, Streamlining temperate phage engineering to facilitate precise in situ manipulation of gut microbiota (1K99GM147604-01). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10507364. Licensed CC0.

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