Microplate computer software

The treated HCC cells (1 × 10⁶) were cultured in 96-well microtiter plates triplicated and incubated at an atmosphere of 5% CO₂ and 37 °C for 5 days. Microplate computer software (Bio-Rad Laboratories, Inc., Hercules, CA, USA) was applied for measuring the OD following the Cell Counting Kit-8 (CCK-8) assay kit protocol (Dojindo, Tokyo, Japan). Then, we plotted the cell proliferation curves. Meanwhile, the cells were treated with ethanol fixation, followed by RNase A treatment and propidium iodide staining. Flow cytometry detection was carried out using FACSCalibur (Becton–Dickinson, Franklin Lakes, NJ, USA) for quantifying cell populations at the G0/G1, S, and G2/M phases, and ModFit software e (Becton–Dickinson) was used. The debris and fixation artifacts of the cells were excluded.

EC9706 cell lines in logarithmic phase growth were seeded into 96-well plates at a density of 1 × 10⁴ cells/well with three replicate wells. After 24 h incubation, when the cells were adhesive, the cells were exposed to a range of concentration of myricetin (0 μM, 25 μM, 50 μM, 100 μM), After 4 hours later, 5-FU (20 μM, 40 μM, 80 μM, 160 μM, 320 μM) was added into the culture. After exposed for 48 h, OD was measured by WST (water-soluble tetrazolium salt) assay using microplate computer software (Bio-Rad Laboratories, USA) according to the protocol of Cell Counting Kit-8 (CCK8) assay kit (Dojindo, Japan). 450 nm absorbance was read on a microplate reader (168–1000 Model 680, Bio-Rad, Hercules, USA). The curves of cell proliferation were plotted. Triplicate parallel experiments were performed for each concentration. The rate of inhibition was calculated by the following equation: rate of growth inhibition (%) = (OD_control – OD_treated)/OD_control × 100%.

The treated HCC cells (1x10⁶) were cultured in 96-well microtiter plates triplicated and incubated at an atmosphere of 5% CO₂ and 37° C for 5 days. Microplate computer software (Bio-Rad Laboratories, Inc., Hercules, CA, USA) was applied for measuring the OD following the Cell Counting Kit-8 (CCK-8) assay kit protocol (Dojindo, Tokyo, Japan). Then, we plotted the cell proliferation curves. Meanwhile, the cells were treated with ethanol fixation, followed by RNase A treatment and propidium iodide staining. Flow cytometry detection was carried out using FACSCalibur (Becton-Dickinson, Franklin Lakes, NJ, USA) for quantifying cell populations at the G0/G1, S, and G2/M phases, and ModFit software e (Becton-Dickinson) was used. The debris and fixation artifacts of the cells were excluded.

The cell viability was monitored using the Cell Counting Kit-8 (CCK8) according to the manufacturer’s protocol. Measured the OD (optical density) value by microplate computer software (Bio-Rad Laboratories, Hercules, CA). Absorbance at 450 nM (A450) was read on a microplate reader (168–1000 Model 680, Bio-Rad).
Colony formation assay was performed to measure the capacity of cell proliferation. Briefly, 1 × 10³ cells were plated in six-well plates. After incubated for 12 days, cells were fixed, stained, photographed, and analyzed.
Tumorigenicity assays in nude mice were performed. Briefly, the mice in groups were inoculated subcutaneously with 1 × 10⁷ cells in the right flank with SNHG7, shSNHG7, or control. Tumor volumes were calculated by using the equation volume (mm³) = A × B²/2, where A is the largest diameter, and B is the perpendicular diameter.