FIG. 7.

Base substitution of synthetic nGFP mRNA reduces but not eliminates its immunogenicity. The cells were transfected with a synthetic nGFP-mRNA (nGFP, 300 ng/mL) that has been modified by base modifications. (A) The transfection was performed twice (1× and 2×) with a 48 h interval. The expression of GFP was exampled with a LSM confocal flourescence microscope at 24 h after each transfection. The boxed area (10T1/2:nGFP, 1×) shows enlarged cells to illustrate the nuclear location of GFP. The morphology of transfected cells was examined under a phase-contrast microscope after TB staining (400×). (B) The mRNA levels of IFNβ, RIG-I, PKR, and TLR3 were determined by RT-qPCR in the cells that were transfected for a second time for 12 h. The results are means±SD of three independent experiments.