Helen Gunter from the BASE mRNA Facility at the University of Queensland in Australia spoke at London Calling 2024 on “Innovations in mRNA vaccine precision manufacturing.” BASE is the largest mRNA manufacturer in Australia, supplying commercial, government, and academic research labs. The plasmid is amplified, linearized, and used as a template to transcribe the mRNA. The mRNA is produced and purified by chromatography. Monitoring every mRNA batch is essential and time-consuming. There is guidance on testing the mRNA integrity and purity. Gunter and the team used Nanopore devices to verify purity. The team was able to detect sub-clonal variants with the RBK114.24 kit. The team also sequenced mRNA to detect the N1 methyl pseudouridine with the Dorado base calling. They used the SQK-RNA004 library kit and sequenced for 24 hours on FLO-MIN004RA. They confirmed the sequence identity for all the mRNAs they analyzed, along with RNA length/integrity. Gunter noted that there was variation in poly(A) tail length. The final product was characterized by a protocol to extract mRNA from the LNP encapsulation for sequencing using direct RNA sequencing library protocols. Gunter was excited about using nanopore sequencing to analyze multivariant vaccines, which can’t be resolved with Sanger sequencing. Multivariant mRNA vaccine sequences can be quantitatively resolved and matched to their input/reference. Double-stranded mRNA and capping analyses are being improved. The application of nanopore sequencing to improve the manufacture of mRNA vaccines offers the speed and resolution needed to ensure clinical manufacturing quality.
