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DNA-Tethered RNA Polymerase for Programmable In vitro Transcription and Molecular Computation

DNA-Tethered RNA Polymerase for Programmable In vitro Transcription and Molecular Computation

Published on: December 29, 2021

A modular cell-free protein biosensor platform using split T7 RNA polymerase.

Megan A McSweeney¹, Alexandra T Patterson¹, Kathryn Loeffler¹

¹School of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, GA 30332, USA.

Science Advances

|February 21, 2025

Summary

This summary is machine-generated.

A new cell-free biosensing platform, T7 RNA polymerase-linked immunosensing assay (TLISA), enables rapid, equipment-free protein detection. This modular technology is suitable for point-of-care diagnostics, offering flexibility and speed.

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Area of Science:

Biotechnology
Molecular Diagnostics
Biosensor Technology

Background:

Conventional protein detection methods are often unsuitable for point-of-care (POC) settings due to cost, complexity, and long development times.
Existing POC assays may have limitations in terms of development speed and adaptability.
There is a need for versatile, equipment-free biosensing platforms for rapid diagnostics.

Purpose of the Study:

To introduce a modular cell-free biosensing platform, T7 RNA polymerase-linked immunosensing assay (TLISA), for generalizable and equipment-free protein detection.
To demonstrate the flexibility and adaptability of the TLISA platform for various protein targets.
To validate TLISA's suitability for point-of-care applications.

Main Methods:

Developed a modular biosensing platform (TLISA) utilizing a split T7 RNA polymerase fused to affinity domains.
Target antigen binding drives polymerase reassembly, initiating reporter gene expression.
Characterized the platform's performance and demonstrated modularity using 16 affinity domains against four different antigens.

Main Results:

TLISA platform demonstrated high modularity with minimal optimization across multiple affinity domains and antigens.
Successfully detected human biomarkers in serum and saliva samples within 1 hour using a colorimetric readout.
The biosensing platform maintained functionality after lyophilization, indicating stability for POC use.

Conclusions:

TLISA offers a highly flexible, modular, and equipment-free approach for protein detection.
The platform is suitable for rapid point-of-care diagnostics, enabling detection in various biological samples.
This technology has the potential to significantly advance diverse protein detection capabilities.