Mimics control

The mimics control, miR-21a mimics, inhibitor control, and anti-miR-21a inhibitor were purchased from GenePharma (GenePharma Co., Ltd., Shanghai). Transfection of miRNA mimics/inhibitors was performed using the Lipofectamine 2000 reagent (Invitrogen) according to the manufacturer’s instruction.

MiR-497 mimics (5′- CAGCAGCACACUGUGGUUUGU-3′, 5′-AAACCACAGUGUGCUGCUGUU-3′), mimics control (5′-UUCUCCGAACGUGUCACGUTT-3′, 5′-ACGUGACACGUUCGGAGAATT-3′), miR-497 inhibitor (5′-ACAAACCACAGUGUGCUGCUG-3′) and inhibitor control (5′-CAGUACUUUUGUGUAGUACAA-3′) were synthesized by Genepharma (Shanghai, China). MiRNAs at 50–100 nM were transfected using Lipofectamine 2000 transfection reagent (Invitrogen, Carlsbad, CA) according to the manufacturer’s protocol.

About 3×10⁵ ZF4 cells were seeded into each well of a 6‐well plate. The next day, dre-miR-100-3p mimic (100 pmol), dre-miR-16b mimic (100 pmol), dre-miR-100-3p inhibitor (300 pmol), dre-miR-16b inhibitor (300 pmol), mimics control (100 pmol) or inhibitor control (300 pmol) (GenePharma, China) was introduced into ZF4 cells respectively using Attractene Transfection Reagent (301005, Qiagen) according to the manufacturer’s instructions. The sequences of above mimics and inhibitors are shown in S1 Table. One day later, ZF4 cells were moved into an incubator at 10°C, 5% CO₂ for cold treatment. After 36 hours, cells were prepared into a cell suspension and stained with 0.4% Trypan Blue solution (w/v) for 5 min, then counted with a hemocytometer.

The mimics control, MiR-138-5p mimics, inhibitor control, and miR-138-5p inhibitor were purchased from GenePharma (GenePharma Co., Ltd., Shanghai). Transfection of miRNA mimics/inhibitors was performed using the Lipofectamine 2000 reagent (Invitrogen) according to the manufacturer’s instruction.

The mimics control, miR-106b mimics, inhibitor control, anti-miR-106b inhibitor, scrambled siRNA control, Tp53inp1 siRNA, and Cdkn1a siRNA were purchased from GenePharma (GenePharma Co., Ltd., Shanghai). Transfection of miRNA mimics/inhibitor or siRNAs was performed using the Lipofectamine 2000 reagent (Invitrogen) according to the manufacturer’s instruction.

To investigate the role of miR-182-5p on GC cell activities, HGC-27 cells in the logarithmic growth phase were transfected with 50 nM miR-182-5p mimics or 8 ng mimics control (GenePharma, Shanghai, China) using 1 μl of the Lipofectamine 2000 reagent. The groups were designed as follows: control group, miR-mimics group, anti-miR group, anti-negative control (NC) group, and miR-NC group. To study the role of RAB27A on GC cell activities, HGC-27 cells were transfected with lentiviral vectors recombined with human RAB27A gene sequence (the constructed vector was plenti-GIII-Ubc-RAB27A). These were assigned to the plenty-RAB27A group and cells transfected with the empty vector plasmid were designated as the NC group (named plenti-Null group).

PcDNA3.1‐HOTAIRM1, pcDNA3.1‐DLGAP1, pcDNA3.1 plasmid vectors, siRNA1‐HOTAIRM1, siRNA2‐HOTAIRM1, miR‐148a mimics, mimics control, miR‐148a inhibitor, and inhibitor control were commercially obtained from GenePharma (Shanghai, China). The sequence of HOTAIRM1 siRNA listed in Table 1. The Fadu cells were plated in 6‐well plates at a density of 1 × 10⁶ per well and cultured in incubator for 24 h at 37°C in 5% CO₂ until the cell confluence arrived 80–90%. Transfections were executed by Lipofectamine 2000 (Invitrogen, Carlsbad, CA) following the instructions of manufacturer. The medium was replaced with complete medium after 6‐hour transfection.