Mir 331 3p mimics

U2932 cells (4×10⁵) in the logarithmic growth phase were seeded into a six-well plate. When cell confluence reached 70–80%, U2932 cells were transfected with pcDNA3.1-UCA1(UCA1 vector, 40 nM), small interfering (si)-UCA1 (40 nM), miR-331-3p mimics (40 nM), miR-331-3p inhibitor (80 nM), or their corresponding negative control (40 nM) by Lipofectamine 2000 (Invitrogen; Thermo Fisher Scientific, Inc.). After transfection at 37°C for 1 h, the cells were incubated with complete medium at 37°C, 5% CO₂ and saturated humidity for another 48 h. Vector plasmid, pcDNA3.1-UCA1, miR-331-3p mimics, miR-331-3p inhibitor and the corresponding NC were obtained from Shanghai GenePharma Co., Ltd.. The sequences were as follows: si-UCA1, 5′-GAGCCGAUCAGACAAACAAUU-3′; si-NC, 5′-UUCUCCGAACGUGUCACGUTT-3′; miR-331-3p mimics, 5′-GCCCCUGGGCCUAUCCUAGAA-3′; miR-331-3p inhibitor, 5′-UUCUAGGAUAGGCCCAGGGGC-3′; mimic NC, 5′-UUCUCCGAACGUGUCACGUTT-3′; inhibitor NC: 5′-CAGUACUUUUGUGUAGUACAA-3′.

Negative control (NC) mimics, miR-331-3p mimics, and miR-inhibitors were designed and synthesized by Genepharm (Shanghai, China). Cells were transfected with NC mimics, miR-331-3p mimics or miR-inhibitors using transfection reagent (pufei biotech, China) following the manufacturer’s instructions.