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Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins

Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins

Published on: October 18, 2022

CRISPRi-based circuits to control gene expression in plants.

Muhammad Adil Khan^1,2, Gabrielle Herring^1,2, Jia Yuan Zhu^1,2

¹Australian Research Council Centre of Excellence in Plant Energy Biology, School of Molecular Sciences, The University of Western Australia, Perth, Western Australia, Australia.

Nature Biotechnology

|May 20, 2024

Summary

This summary is machine-generated.

Researchers developed a CRISPR interference (CRISPRi) gene circuit platform for plants, enabling precise control over gene expression. This modular system offers programmability and reversibility for synthetic biology applications in diverse plant species.

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Area of Science:

Synthetic Biology
Plant Biotechnology
Molecular Biology

Background:

Synthetic gene circuit construction in plants is hindered by a scarcity of adaptable and independent components.
Existing methods lack the orthogonality and modularity required for complex genetic programming.

Purpose of the Study:

To establish a CRISPR interference (CRISPRi)-based, reversible gene circuit platform in plants.
To engineer a toolkit of repressible promoters and logic gates for sophisticated gene expression control.

Main Methods:

Implementation of a CRISPRi system with engineered repressible promoters in Arabidopsis thaliana protoplasts.
Development of NOT and NOR logic gates and optimization of single-guide RNA expression.
Testing of CRISPRi gate functionality in stably transformed Arabidopsis and cross-species protoplasts (Physcomitrium patens, Triticum aestivum, Brassica napus).

Main Results:

Successful construction and demonstration of NOR gates in Arabidopsis protoplasts and whole plants, showcasing programmability and reversibility.
Validation of CRISPRi logic gate activity across multiple plant species.
Creation of OR, NIMPLY, and AND logic functions by integrating multiple NOR gates, confirming system modularity.

Conclusions:

The developed CRISPRi platform provides orthogonal, compact, reversible, programmable, and modular synthetic gene circuits for plants.
This system enables sophisticated spatiotemporal control of gene expression, advancing plant synthetic biology.