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Related Experiment Video

Updated: May 3, 2026

Determination of S-Phase Duration Using 5-Ethynyl-2'-deoxyuridine Incorporation in Saccharomyces cerevisiae

Determination of S-Phase Duration Using 5-Ethynyl-2'-deoxyuridine Incorporation in Saccharomyces cerevisiae

Published on: October 21, 2022

An orthogonal DNA replication system in yeast.

Arjun Ravikumar¹, Adrian Arrieta¹, Chang C Liu²

¹Department of Biomedical Engineering, University of California-Irvine, Irvine, California, USA.

Nature Chemical Biology

|February 4, 2014

Summary

This summary is machine-generated.

Researchers created a novel extranuclear DNA replication system in yeast using engineered DNA polymerases. This system allows for targeted mutations on plasmids without affecting the host genome, enabling controlled evolution studies.

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Genetic Studies of Human DNA Repair Proteins Using Yeast as a Model System

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Published on: March 18, 2010

Area of Science:

Synthetic biology
Molecular and cell biology
Biotechnology

Background:

Developing controllable DNA replication systems is crucial for genetic engineering and synthetic biology.
Existing methods often lack specificity or impact the host genome.
Targeted mutagenesis requires precise control over DNA replication and repair mechanisms.

Purpose of the Study:

To engineer an orthogonal DNA replication system in Saccharomyces cerevisiae.
To demonstrate targeted, in vivo mutagenesis using engineered error-prone DNA polymerases.
To establish a platform for in vivo continuous evolution independent of host genomic constraints.

Main Methods:

Construction of an orthogonal DNA plasmid-DNA polymerase replication system.
Engineering of error-prone DNA polymerases for targeted mutagenesis.
Culturing of Saccharomyces cerevisiae with the engineered system.
Quantification of mutation rates on plasmids and genomic DNA.

Main Results:

Successful establishment of an extranuclear replication system in yeast.
Engineered polymerases achieved substantial, stable increases in per-base mutation rates on the plasmid.
No significant increase in mutation rates was observed in the host genome.
Demonstrated mutational targeting in vivo.

Conclusions:

The developed orthogonal replication system enables precise control over plasmid mutations.
This system serves as a versatile platform for in vivo continuous evolution.
The ability to manipulate replicative properties independently of the host opens new avenues in genetic engineering.