SARS-CoV-2 S/N Protein Expression Plasmid

The SARS-CoV-2 S/N protein-encoding gene, containing an N-terminal Kozak sequence (GCCACC) followed by an initiation codon (ATG), was synthesized using a mammalian-optimized codon (GenScript Co., Nanjing, China). It was then cloned into the expression vector pcDNA3.1 (+) via EcoRI and XbaI digestion and named pS/pN (DNA vaccines) (Figure 1A). The pE/pM protein was constructed and identified as described previously [15 (link)]. Vaccines were prepared using endotoxin-free Maxiprep kits (Qiagen, Beijing, China), and sequences were confirmed using Sanger DNA sequencing. The expression of the S/N protein was confirmed using western blotting and anti-S (Sino Biological, Beijing, China)/anti-N antibodies diluted at 1:1000. These experiments were conducted as described previously [15 (link),18 (link)].