Scrambled sirna control si nc

Small interfering RNA (siRNA) against XIST (si-XIST) or PAPPA (si-PAPPA) and siRNA scrambled control (si-NC), XIST overexpression vector (pcDNA-XIST), and its negative control (pcDNA) were acquired from GenePharma (Shanghai, China). MiR-98-5p mimic (miR-98-5p) and control (miR-NC) and miR-98-5p inhibitor (anti-miR-98-5p) and control (anti-NC) were provided by Sangon (Shanghai, China). Different oligonucleotides or vectors were transfected into HUVECs with Lipofectamine 2000 reagent (Qiagen) following the user’s guidebook.

Specific small interfering RNA (siRNA) against circHIPK3 (si-circHIPK3) and siRNA scrambled control (si-NC), and short hairpin RNA objecting circHIPK3 (sh-circHIPK3) and shRNA scrambled control (sh-NC) were purchased from GenePharma (Shanghai, China). The miR-876-5p mimic (miR-876-5p) and its negative control (miR-NC), and miR-876-5p inhibitor (anti-miR-876-5p) and its negative control (anti-miR-NC) were designed and acquired from Thermo Fisher Scientific (Waltham, MA, USA). HGC-27 and AGS cells were transfected with plasmids or oligonucleotides using Lipofecamine2000 (Invitrogen, Carlsbad, CA, USA). The sequences for si-circHIPK3, si-NC, sh-circHIPK3 and sh-NC were as follows: si-circHIPK3: 5′-CUACAGGUAUGGCCUCACA-3′, si-NC: 5′-UUCUCCGAACGUGUCACGUTT-3′, sh-circHIPK3: 5′-ccggCUACAGGUAUGGCCUCACAttcaagagaTGTGAGGCCATACCTGTAGTTTTTTGGTACC-3′, sh-NC: 5′- ccggUUCUCCGAACGUGUCACGUTTttcaagagaAATCGTGACACGTTCGGAGAATTTTTTGGTACC-3′. For overexpression of PIK3R1 (PIK3R1), the primers were used for amplification and then cloned in the mammalian expression pcDNA3.1 vector (Invitrogen), PIK3R1 F-5′-CCGGAATTCATGAGTGCTGAGGGGTACCAGTAC-3′; and R-5′-CCGCTCGAGATCGCCTCTGCTGTGCATATATA-3′.

Specific small interfering RNA (siRNA) against circPOSTN (si-circPOSTN) or against TPX2 (si-TPX2) and siRNA scrambled control (si-NC), plasmid-mediated pcDNA and pcDNA-circPOSTN, miR-361-5p mimic and miR-NC, anti-miR-361-5p and anti-miR-NC were purchased from Genepharma (Shanghai, China). The lentiviral vectors of specific short hairpin RNA (shRNA) target circPOSTN (sh-circPOSTN) and shRNA scrambled control (sh-NC) were constructed by Sangon (Shanghai, China). For transfection assay, LN229 and U251 cells were seeded into 6-well plates (3 × 10⁴ cells/well) overnight, and cell density reached 70–80% prior to transfection. The transfection analysis was conducted with Lipofectamine 2000 (Invitrogen) according to the recommendations. In addition, the concentrations of oligonucleotides for transfection were 50 nM. The concentrations of plasmids for transfection were 100 nM.