Improving mRNA vaccine manufacture using RNA sequencing

The manufacture mRNA vaccines needed to manufacture billions of doses at sufficient quality during the COVID19 pandemic. The removal of contaminating RNAs and the inclusion of modified nucleotides reduces the innate immune response and improves the translation and performance of mRNA vaccines and therapies. Here we validate the use of nanopore sequencing during the manufacture and analysis of mRNA vaccines. Using a modified protocol and software toolkit, we show how nanopore sequencing enables quantitative analysis of mRNA vaccine sequence, length and purity, including off target RNA impurities. These analyses inform best practise approaches for designing mRNA sequences, optimise manufacturing processes and benchmark purification methods. We also perform direct RNA sequencing of mRNA vaccines to measure pseudouridines incorporation errors and 5 prime capping. Together this shows how nanopore sequencing can be used during mRNA manufacture to validate batches, monitor quality and provide a detailed characterisation of the final mRNA vaccine product.