# Synthetic genetic systems for rapid biomolecular evolution in vivo

> **NIH NIH R35** · UNIVERSITY OF CALIFORNIA-IRVINE · 2024 · $442,016

## Abstract

Project Summary/Abstract
Over the past five years, my lab has made significant strides in the development of genetic systems capable of
driving the rapid mutation and evolution of user-selected genes of interest in vivo. These systems have allowed
us and others to quickly and scalably evolve enzymes, proteins, and antibodies to address a range of problems
spanning from studying drug resistance to creating affinity reagents on demand. These systems have also
begun to allow us to use rapid mutational accumulation as a method for tracing cell lineage in developing
animals. One of our key accomplishments has been the invention of an orthogonal DNA replication system
(OrthoRep). In OrthoRep, an error-prone orthogonal DNA polymerase (DNAP) exclusively replicates a special
cytosolic plasmid encoding only genes of interest (GOIs), driving their continuous evolution fully in vivo. This
MIRA will integrate our lab’s work on OrthoRep and support its further development and application in the next
five years. In particular, we will grow the core OrthoRep technology in order to accelerate GOI evolution in
yeast even more than we currently have, attempt to establish OrthoRep in mammalian cells in order to extend
the range of problems OrthoRep can directly address, apply OrthoRep to the engineering of bespoke Cas9s to
extend the range and efficacy of targeting, apply OrthoRep to improve the capabilities of a lineage tracing tool
developed by my lab, and apply OrthoRep to the generation of mutually orthogonal collections of aminoacyl-
tRNA synthetase (aaRS)/tRNA pairs to support genetic code expansion efforts also ongoing in my lab.
We
hope the set of activities proposed for this MIRA will solidify OrthoRep as an exceptionally powerful genetic
system for evolving enzymes and proteins capable of solving high-reward problems in the chemical, biological,
and biomedical sciences.

## Key facts

- **NIH application ID:** 10839474
- **Project number:** 5R35GM136297-05
- **Recipient organization:** UNIVERSITY OF CALIFORNIA-IRVINE
- **Principal Investigator:** Chang C Liu
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $442,016
- **Award type:** 5
- **Project period:** 2020-06-01 → 2026-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10839474

## Citation

> US National Institutes of Health, RePORTER application 10839474, Synthetic genetic systems for rapid biomolecular evolution in vivo (5R35GM136297-05). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10839474. Licensed CC0.

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