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. 2020 Jul 21;32(3):107920. doi: 10.1016/j.celrep.2020.107920

Figure 2.

Figure 2

Construction and Recovery of a Chimeric Lyssavirus G Vaccine

(A) Viral genome schematics. BNSP333 is the parent vaccine vector based on RABV strain SAD B19. Its G is located in the native fourth position and contains the attenuating R333E mutation. BNSPΔG is based on BNSP333 but lacks the native G. All of the following experimental constructs are based on BNSPΔG: rRABV contains a human codon-optimized (c.o.) RABV G with the attenuating mutation R333E at the second position; rMOKV contains human c.o. MOKV G at the second position; rChimera1 (LyssaVax) contains Chimeric G 1, with the attenuating R333E mutation, at the second position; and rChimera2 contains Chimeric G 2 at the second position.

(B) Infection immunofluorescence. VERO cells infected with either LyssaVax (left column), rMOKV (second column), rRABV (third column), or uninfected (right column) were fixed and stained with a DyLight 488-conjugated human anti-RABV G mAb 4C12 (green) and mouse anti-MOKV G sera (red). Nuclei labeled in blue by DAPI. Scale bars represent 50 μm.

(C) Analysis of purified virions. 3-μg viral particles denatured and resolved by SDS-PAGE, then total protein stained with SYPRO Ruby. Viral proteins are indicated.