Engineering circular RNA for potent and stable translation in eukaryotic cells

Area of Science:

• Molecular Biology
• Biotechnology
• Biochemistry

Background:

• Messenger RNA (mRNA) is crucial for biological applications but limited by its short half-life.
• Extending the stability and protein production duration of RNA is a key challenge in biotechnology.

Purpose of the Study:

• To develop exogenous circular RNA (circRNA) as a method for prolonged protein expression.
• To engineer and optimize circRNA for efficient protein synthesis in eukaryotic cells.

Main Methods:

• Engineered a self-splicing intron for efficient in vitro circularization of RNAs up to 5 kb.
• Designed accessory sequences to facilitate splicing and maximize translation from circRNAs.
• Purified engineered circRNA using high-performance liquid chromatography (HPLC).

Main Results:

• Demonstrated efficient circularization of diverse RNA molecules.
• Achieved robust and stable protein production from engineered circRNAs in eukaryotic cells.
• HPLC-purified circRNA showed superior protein production quantity and stability.

Conclusions:

• Exogenous circRNA serves as a powerful tool for sustained protein expression.
• CircRNA represents a promising alternative to linear mRNA for biotechnological applications.
• This study establishes circRNA as a viable platform for enhanced protein production.