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. 2019 Jan 31;8:F1000 Faculty Rev-134. [Version 1] doi: 10.12688/f1000research.16989.1

Figure 2. Generation of r3ML29.

Figure 2.

BHK-21 cells were transfected with pol-II expression plasmids for ML29 L and NP, required to support viral replication and transcription, together with plasmids that direct pol-I-mediated intracellular synthesis of L, and recombinant S1 and S2 RNA genome species. Six days later, tissue culture supernatants were collected and used to infect Vero cells to amplify the rescued r3ML29 that were plaque-purified and scaled up to generate viral stocks. ( A) Genome organization of r3ML29/eGFP. ( B) r3ML29/EGFP grows to high titers in Vero cells. Cells were infected (multiplicity of infection = 0.1) and virus titers in tissue culture were determined at the indicated times. The rML29 reverse genetics is an advanced vaccine platform to further improve the safety and immunogenicity of Lassa virus vaccine candidates. FFU, focus-forming units; GFP, green fluorescent protein; GPC, glycoprotein precursor; h.p.i., hours post-infection; LoD, limit of detection; NP, nucleoprotein; TCS, tissue culture supernatant.