Development of Candidate Vaccine Viruses for Influenza Using Reverse Genetic Technology

Begin with a mixture of eight plasmids to generate a candidate vaccine virus (CVV), a prototype influenza virus used in vaccine production.

Each plasmid carries a complementary DNA (cDNA) copy encoding one gene segment of the influenza virus’s segmented genome.

Six plasmids encode internal genes, and two encode antigen genes engineered to reduce viral pathogenicity.

Add the plasmid mixture to a suspension of mammalian epithelial cells.

Deliver a short electric pulse to induce temporary membrane pores, enabling plasmid uptake.

Inside the nucleus, each plasmid is transcribed into a viral RNA for genome replication and an mRNA.

The internal gene mRNAs encode proteins for viral replication and assembly.

The mRNAs from the antigen genes encode surface antigens.

The viral components assemble and bud from the cells, forming CVVs that display engineered surface antigens.

The CVVs are now ready for further amplification.