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In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity

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Programmable in vivo selection of arbitrary DNA sequences.

Tuval Ben Yehezkel¹, Tamir Biezuner, Gregory Linshiz

¹Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot, Israel.

|November 17, 2012

Summary

Synthetic DNA devices can be programmed to interface with bacterial DNA mismatch repair systems for function-independent DNA selection in vivo. This breakthrough enables programmable selection of DNA variants within living cells.

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Published on: March 29, 2019

Area of Science:

Synthetic Biology
Molecular Biology
Genetics

Background:

Natural intracellular machinery possesses high fidelity and regulatory capacity within cells.
Synthetic biomolecular components can be engineered to interact with cellular transcription, splicing, and translation processes.
Interfacing synthetic components with intracellular machinery in vivo is crucial for harnessing cellular processing power.

Purpose of the Study:

To demonstrate programmable, function-independent DNA selection within a living bacterial system.
To engineer synthetic DNA devices that interface with endogenous cellular machinery.
To establish a logical-functional interface between synthetic components and cellular systems.

Main Methods:

Designed a library of synthetic DNA devices with input DNA sequences and logical selection modules.
Engineered devices to interface with the bacterial DNA mismatch repair (MMR) system in vivo.
Utilized the MMR system to select for the most abundant DNA variant, irrespective of its function.

Main Results:

Successfully demonstrated programmable, function-independent DNA selection in vivo.
Developed synthetic DNA devices capable of interacting with the bacterial MMR system.
Showcased a novel logical-functional interface between engineered synthetic components and a complex endogenous cellular system.

Conclusions:

The engineered synthetic DNA devices provide a proof of concept for programmable DNA selection in vivo.
This work presents a unique interface between synthetic biology and endogenous cellular repair mechanisms.
Further research may lead to new functional devices for diverse cellular processes in research and applications.