Reporter vector system

To confirm that DLEU1 and PRPS1 were targets of miR-320b, LncRNA DLEU1-WT (containing the binding sites of miR-320b at DLEU1), LncRNA DLEU1-Mut (mutation of binding sites), PRPS1-WT (containing the binding sites of miR-320b at PRPS1 3’UTR) and PRPS1-MUT (mutation of binding sites) were cloned into Luciferase Reporter Vector (Invitrogen, CA, USA). LncRNA DLEU1-WT, LncRNA DLEU1-Mut, PFKL-WT or PFKL-MUT were co-transfected with miR-320b mimics into LoVo and SW480 cells. After 48-h transfection, LoVo and SW480 were explored using a Reporter Vector System (Invitrogen, CA, USA) using a Glomax20/20 luminometer (Promega, WI, USA) (27 (link)).

PFKL-WT (containing the binding sites of miR-185-3p at PFKL 3′-UTR) and PFKL-MUT (mutation of binding sites) were cloned into Luciferase Reporter Vector (AM5795, Invitrogen, CA, United States). PFKL-WT or PFKL-MUT was co-transfected with miR-185-3p mimic into H1299/ER or A549/ER cells. After transfection for 48 h, H1299/ER and A549/ER cells were evaluated using the Reporter Vector System (AM5795, Invitrogen, CA, United States) using a GloMax 20/20 Luminometer (Promega, WI, United States) (Zhang et al., 2020 ).

In order to confirm that WT1 was a target of rno-miR-128-3p, WT1-WT (containing the binding sites of rno-miR-128-3p at WT1 3′UTR) and WT1-MUT (mutation of binding sites) were cloned into Luciferase Reporter Vector (AM5795, Invitrogen, Carlsbad,, CA). WT1-WT or WT1-MUT were co-transfected with rno-miR-128-3p mimics into GCs. Forty-eight hours after transfection, GCs were explored using a Reporter Vector System (AM5795, Invitrogen, CA) via a Glomax20/20 luminometer (Promega, Madison, WI) (Zhang et al. 2017b (link)).