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Updated: Jan 17, 2026

Large-scale Production of Recombinant RNAs on a Circular Scaffold Using a Viroid-derived System in Escherichia coli

Large-scale Production of Recombinant RNAs on a Circular Scaffold Using a Viroid-derived System in Escherichia coli

Published on: November 30, 2018

Purifying circular RNA by ultrafiltration.

Karen Guillen-Cuevas¹, Xiaoming Lu¹, Marc R Birtwistle¹

¹Department of Chemical and Biomolecular Engineering, Clemson University, 127 Earle Hall, 206 S. Palmetto Blvd., Clemson, SC 29634 USA.

Separation and Purification Technology

|September 19, 2025

Summary

This summary is machine-generated.

Ultrafiltration effectively purifies circular RNA (circRNA), overcoming limitations of linear messenger RNA (mRNA). This method achieves high purity and yield, paving the way for scalable circRNA therapeutic manufacturing.

Keywords:

RNA gene expression RNA production RNA purification methods circular RNA therapeutics therapeutic RNA

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Last Updated: Jan 17, 2026

Large-scale Production of Recombinant RNAs on a Circular Scaffold Using a Viroid-derived System in Escherichia coli

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Published on: November 30, 2018

Fecal micro RNA Isolation

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Identification of Circular RNAs using RNA Sequencing

Identification of Circular RNAs using RNA Sequencing

Published on: November 14, 2019

Area of Science:

Biotechnology
Molecular Biology
Biochemistry

Background:

Messenger RNA (mRNA) vaccines have spurred interest in mRNA therapeutics.
Linear mRNA is unstable due to enzymatic degradation, unlike more stable circular RNA (circRNA).
Current methods struggle to purify circRNA from linear contaminants produced during in vitro transcription (IVT).

Purpose of the Study:

To investigate ultrafiltration for purifying protein-encoding circRNA from IVT reactions.
To assess the feasibility of using polyethersulfone membranes for circRNA separation.
To determine optimal operating conditions for circRNA purification via ultrafiltration.

Main Methods:

Measured sieving coefficients of circRNA, linear precursor RNA, and nicked RNA using membranes with varying molecular weight cutoffs (30-300 kDa).
Analyzed separation performance based on permeate flux.
Estimated critical fluxes for RNA separation and identified suitable operating parameters.

Main Results:

Achieved 86% purity and >50% yield for circRNA using ultrafiltration.
Demonstrated superior performance compared to size-exclusion high-performance liquid chromatography (SE-HPLC), which yielded 41% purity and 45% yield.
Identified optimal ultrafiltration conditions for circRNA purification.

Conclusions:

Ultrafiltration is a highly effective and superior method for purifying circRNA at the research scale.
The scalability of ultrafiltration in bioprocessing supports its potential for large-scale circRNA therapeutic manufacturing.
This study provides a foundation for advancing circRNA-based therapeutics through improved purification techniques.