Escherichia coli s30 system

The cell-free bacterial translation and mammalian translation assays were performed by the commercially available Escherichia coli S30 System and Rabbit Reticulocyte Lysate System (Promega), respectively. The assays were performed as described by the manufacturer, in conjunction with appropriate positive control (chloramphenicol) and negative control (ampicillin) antibiotics. In bacterial translation assay, the reaction mixtures were incubated at 37°C for 1 h. Mammalian translation assay reaction mixtures were incubated at 30°C for 1 h. Luciferase assay reagent was added to the reaction and the intensity of the luminescence was measured by luminescence microplate reader (FLx800 BioTek Instruments, Inc. Winooski, Vermont) according to the manufacturer’s instructions. Average luciferase readout of protein production from two replicates from two independent experiments was calculated.

The cell-free bacterial transcription/ translation assay was performed using Escherichia coli S30 System (Promega). The assay was carried out as per the manufacturer’s instructions. Gentamicin was used as a positive control. Briefly, simvastatin and gentamicin were added at the indicated concentrations to the reaction mixtures and incubated at 37 °C for one hour. The intensity of luminescence was quantified using a standard FLx800 microplate reader (BioTek Instruments, Inc. Winooski, Vermont) after addition of the luciferase assay reagent.