Advanced Plasmid Linearization Using Type I And Type II Restriction Enzymes: A Scalable, High-Purity Platform For Cell And Gene Therapy Applications

By Charlie Fan; Delicia Henriques, PhD; Justin Chen; Philemon Asfeha; Rui Tang; Pawan Bhatt

Plasmid DNA is a foundational material for cell and gene therapies, powering applications from viral vector production to mRNA vaccines and CAR-T treatments. While supercoiled plasmids are widely used, linearization offers distinct advantages for certain processes, including enhanced transfection efficiency, improved transcription accessibility, and stronger gene expression in non-viral delivery systems.

This approach uses restriction enzymes to convert circular plasmids into linear forms, ensuring structural precision and compatibility with advanced therapeutic workflows. Recent developments demonstrate a robust platform that combines high-purity plasmid production with efficient linearization, achieving over 92% linearized DNA and endotoxin levels below 10 EU/mg. The process delivers consistent results across multiple plasmid types, with recovery rates exceeding 40% and host cell protein impurities under 1%.

With a streamlined timeline of approximately four weeks from strain development to final QC release, this method supports scalability and regulatory compliance for clinical and commercial manufacturing. Explore how optimized linearization strategies are accelerating next-generation therapies and improving outcomes in gene delivery.