Table 1.

Comparison of Two Investigations into the Immunogenicity of Exogenous CircRNAs

Experiment	“N6-Methyladenosine Modification Controls Circular RNA Immunity” [52]	“RNA Circularization Diminishes Immunogenicity and Can Extend Translation Duration In Vivo” [53]
Type of linear RNA compared to circRNA	5’ hydroxyl linear RNA	5’ triphosphate linear RNA
RNase R treatment of in vitro transcribed circRNA	2–3 hours	15 min
Phosphatase treatment	2 hours	15 min
Experiment readout	Measure immune gene expression	Measure cytokine and chemokine in culture medium
RNA transfection	Forward, used Lipofectamine 2000	Reverse, used Lipofectamine MessengerMax
RNA transfection efficiency	Measured levels of RNA delivered for normalization	Did not measure levels of RNA delivered