Pcdna snhg15

The siRNAs specifically targeting SNHG15 (si-SNHG15–1, si-SNHG15–2 and si-SNHG15–3), scrambled negative control (si-control), pcDNA-SNHG15, empty pcDNA vector (vector), miR-141 mimics (miR-141), mimics negative control (miR-control), anti-miR-141 and anti-miR-control were commercially synthesized by Genepharma (Shanghai, China). For transient transfection, U2OS and MG63 cells were plated into six-well plates (2 × 10⁵/well) and routinely maintained for 24 h at 37 °C. Then the cells were transfected with siRNAs, pcDNA-SNHG15, vector, miR-141, miR-control, anti-miR-141, anti-miR-control, si-SNHG15 + anti-miR-141, si-SNHG15 + anti-miR-control, pcDNA-SNHG15 + miR-141, or pcDNA-SNHG15 + miR-control by Lipofectamine 2000 (Invitrogen) according to the manufacturer’s protocol. Subsequent experiments were performed at 48 h post transfection. The sequences of si-RNAs, anti-miR-141 are as follows: si-SNHG15–1 (Sense: 5′-CAG GTA GAC CGT GCA CGT AA-3′, Anti-sence:3′-CCT TGA TGC GTT GCC AGC AGA-5′), si-SNHG15–2 (Sense: 5′-CCG TGC GTA AAC GTT TGC CA-3′, Anti-sence: 3′-TGG CGG TAA CGT AAA TGC G-5′), si-SNHG15–3 (Sense: 5′-ACG GTG GCA ACG TGC GTG GCC A-3′, Anti-sence: 3′-GCC TGC AAC GGT GCA AAT GCG-5′), anti-miR-141 (CCA UCU UUA CCA GAC AGU GU UA).

Small interfering RNA (siRNA) negative control (si-NC), siRNA against SNHG15 (si-SNHG15), siRNA against GFRA1 (si-GFRA1), pcDNA, and SNHG15 overexpression plasmid (pcDNA-SNHG15) were purchased from GenePharma (Shanghai, China). Anti-miRNA-NC (anti-miR-NC), anti-miR-381-3p, miR-NC, and miR-381-3p were synthesized by RiboBio (Guangzhou, China). All transfection experiments were conducted using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA).