The pmiRGLO vectors containing wild-type (WT) or mutant (MUT) GCNT4 3ʹ-UTRs sequences were purchased from Genechem (Shanghai, China). The MKN45 and AGS cells were treated with 0.24 µg of either vector and 40 nM ofmiR-130a-3p or miR-130a-3p NC using Lipofectamine 3000. After 72 h of transfection, the Luciferase Assay Kit (Cat#: RG027; Beyotime, China) was used for dual-luciferase activity measurements. The final results were represented by firefly luciferase activities relative to that of Renilla luciferase [22 (link),25 (link)].
Pmirglo vector
Manufactured by Genechem
Sourced in China
The PmiRGLO vectors are a set of plasmid-based tools designed for the study of microRNA (miRNA) function. These vectors allow for the expression and detection of miRNAs in various cell lines and model systems. The core function of the PmiRGLO vectors is to facilitate the investigation of miRNA-mediated gene regulation through reporter-based assays.
Automatically generated - may contain errors
Lab products found in correlation
Luciferase assay kit, by Beyotime (2 mentions)
Dual luciferase reporter assay system, by Promega (2 mentions)
Dual luciferase reporter assay kit, by Promega (1 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
Dual luciferase assay system, by Promega (1 mentions)
Site direct mutagenesis kit, by Takara Bio (1 mentions)
6 protocols using pmirglo vector
1
Dual-Luciferase Assay for GCNT4 3'UTR Regulation
2
Luciferase Assay for LRP8 3'UTR
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The pmiRGLO vectors containing LRP8 3′-UTRs wild type (WT) or mutant (MUT) sequences were purchased from Genechem (Shanghai, China). Approximately 5 × 103 CaOV3 and SKOV3 cells were seeded in 24-well plates overnight. Next, the cells were treated with pmiRGLO LRP8 3′-UTR WT, MUT1, or MUT2 plasmids, and miR-362-3p mimic or NC was used to treat the cells using Lipo3000. After 48 h, the luciferase assay kit (Cat#: RG027; Beyotime, China) was used to examine the relative luciferase activity normalized to renal luciferase activity.
3
Dual-Luciferase Assay for CircRNA-miRNA Interaction
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GeneChem constructed the pmir-Glo vectors containing hsa_circ_0000285 (WT-hsa_circ_000028/Mut-hsa_-circ_0000285) and CCNB2 30 UTR (WT-CCNB2/Mut-CCNB2) mutant (Mut) or wild-type (WT) sequences that could bind to miR-582-3p. The mimic-NC and miR-582-3p mimic were also obtained from GeneChem. With the aid of a Lipofectamine 2000 Transfection Reagent, a combination of a vector and either the mimic-NC or miR-582-3p mimic was introduced into the Huh7 and Hep 3B cells. Following 48-h incubation, a dual-luciferase reporter assay system (Promega, Madison, WI, USA) was utilized to assess the luciferase activities in the transfected cells. 26 (link)
4
Luciferase Reporter Assay for FOXD1 Variants
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FOXD1‐AS1‐WT/MUT and FOXD1‐WT/MUT were established by GenePharma and sub‐cloned into the pmirGLO vector (GeneChem). FOXD1‐AS1‐WT/MUT or FOXD1‐WT/MUT was transfected into HKT‐293T cells with indicated transfection plasmids. Luciferase activity was tested by applying Dual‐Luciferase Reporter Assay Kit (Promega, Madison, WI, USA).
5
Identification of XPO1 as miR-483-3p Target
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The wild‐type (Wt) and mutant (Mut) binding site of XPO1 3′ UTR was respectively subcloned into pmirGLO vector (GeneChem) to construct XPO1‐Wt/Mut. Then, XPO1‐Wt or XPO1‐Mut was cotransfected with NC mimics or miR‐483‐3p mimics into PC12 cells using Lipofectamine 2000 (Invitrogen). The luciferase activity was evaluated by a Dual‐Luciferase Assay System (Promega, Madison, WI, USA) 48 h after plasmid transfection.
6
Luciferase Reporter Assay for miRNA Targets
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The amplified wild-type FOXP4-AS1 or CBX4 sequences were inserted into pmirGLO vector (GeneChem, Shanghai, China) to generate FOXP4-AS1-wt or CBX4-wt. Site-direct mutagenesis kit (Takara) was used to generate mutant luciferase vectors and termed as FOXP4-AS1-mt or CBX4-mt. Cells were co-transfected with luciferase constructs and miR-136-5p mimic or NC-mimic using Lipofectamine 2000. Dual-Luciferase Reporter Assay System (Promega, Madison, WI, USA) was used to measure relative luciferase activity after 48 h of transfection using Renilla luciferase as internal control. Experiments were repeated in triplicates.
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