Schematic of potential outputs from bicistronic IRES reporter. The bicistronic reporter comprises a vector that makes an mRNA with a firefly luciferase (yellow) followed by a stop codon, followed by an IRES (green), followed by a renilla luciferase (orange). In the true positive scenario, cap-dependent translation drives translation of the firefly luciferase and the IRES drives translation of the renilla luciferase. A false positive result can occur when a cryptic promoter (gray) can create a new mRNA where the cap can drive translation of the renilla luciferase. A false positive result can also occur when cryptic splicing (gray) can splice out the IRES and stop codon and cause the renilla to be translated through a cap-dependent mechanism.
