Endogenous mature mirna mimics

The endogenous mature miRNA mimics, inhibitors, and antagomirs were purchased from RiboBio (Guangzhou, China). miRNA mimics, miRNA inhibitors, and miRNA NC were transfected into cells using Lipofectamine 2000 (Invitrogen, Carlsbad, USA), according to the manufacturer’s instructions. After 48?h of transfection, the cells were used for further experiments. The pDonR223-WNT5A plasmid carrying the human WNT5A (long isoform) gene was purchased from Axybio Bio-Tech Co., Ltd (Changsha, China). The complete coding sequence of human WNT5A (long isoform) was amplified from the pDonR223-WNT5A plasmid. PCR products representing the WNT5A (long isoform) coding sequence and pEGFP-N1 plasmid were digested with XhoI and HindIII, and the fragments were purified and ligated with T4 DNA ligase. The ligated product was transformed into Top10 competent cells, and the positive clone was referred to as pEGFP-N1-WNT5A. The oligonucleotides targeting WNT5A and corresponding cholesterol and methylation modified scramble sequences (5’-CATGACATCAGTCGGAGAA-3’) was obtained from RiboBio (RiboBio Co. Ltd., China) and transfected into 1299 cells according to the manufacturer’s protocols.

All endogenous mature miRNA mimics, inhibitors, and agomirs were purchased from RiboBio (Guangzhou, China). For transfection, experimental protocols were performed according to the manufacturer’s instructions (RiboBio). The miRNA mimics, miRNA inhibitors, and miRNA NC were transfected into cells using Lipofectamine 2000 (Invitrogen, Carlsbad, USA) according to the manufacturer’s instructions. After a 48-h transfection, the cells were used for further experiments.

All endogenous mature miRNA mimics, inhibitors and antagomirs were purchased from RiboBio (Guangzhou, China). Transient transfections of miRNA mimics/antagomirs/inhibitors were carried out using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s protocol. Six hours following transfection, transfection media were replaced with culture media. All miRNA transfections were for 48 hours.