Figure 1

Strategy to produce DENV-2 EDIII containing VLPs in P. pastoris . (A) Map of pAO815-based P. pastoris-integrative expression vector carrying HBsAg (S) and EDIII-2-HBsAg (ES) antigen gene expression cassettes in tandem. Each gene is present in an independent expression cassette with the AOX1 promoter (P) on the 5’ side and the AOX1 transcription terminator (T) on the 3’ side. A family of vectors was created in which the number (n) of S gene expression cassettes varied from 0–4. Other elements present in the vector include the ampicillin resistance marker (Amp), the bacterial replication origin (Ori), 3’ terminal sequences of the AOX1 gene (3’AOX1) and the yeast histidinol dehydrogenase marker gene (HIS4). The dashed arrows point to symbols used to depict the proteins encoded by the ‘S’ (red sphere) and ‘ES’ (red and blue sphere) genes. Unique sites used for cassette assembly are shown. (B) Schematic representations of the putative VLPs predicted to be generated from the construct when the ‘S’ antigen copy number is 0 (ES,S₀), 1 (ES,S₁), 2 (ES,S₂) or 4 (ES,S₄) per copy of the ‘ES’ fusion antigen.