Fig. 3

Introduction of enhanced green fluorescent protein (EGFP) mRNA expression in methionine-deficient medium to evaluate HeLa cell viability. A EGFP mRNA levels were quantified using quantitative real-time polymerase chain reaction (qRT-PCR) and normalized to GAPDH from endogenous human initiator tRNA (dark violet) in HeLa cells. Samples of EGFP-positive cells in an L-methionine (L-Met)–deficient medium (orange), HeLa cells only (red), and endogenous human RNAi were expressed in methionine-positive HeLa cells (dark violet). The EGFP expression levels are shown as the relative fluorescence populations in the dot plots. A flow cytometry analysis was conducted using EGFP cells in the absence or presence of L-Met, as follows: HeLa cells only (red), L-Met positive media (dark violet), and human initiator tRNA in an L-Met-free medium (orange). All data obtained in the FACS and qRT-PCR experiments are expressed as the mean ± standard deviation of three independent experiments. B Effect of L-Met on HeLa cell viability. HeLa cells were treated with L-Met (red bar) or without L-Met (orange bar) for 24 h. Cell viability was determined using CCK8 assays and calculated relative to that of the control HeLa cells (L-Met sufficiency). Data are presented as the mean ± standard error of the mean (SEM) for three independent experiments