Wild‐type (wt) or mutant (mt) promoter sequence of BRCA1 or lnc‐HZ10 was fused into the luciferase pmirGLO‐basic reporter vector (Madison, Promega, USA). Swan 71 cells were co‐transfected with 300 ng pmirGLO‐wt/mt‐BRCA1 or pmirGLO‐wt/mt‐HZ10 and 200 nm AhR plasmid or its vector in Lipofectamine 3000 (Invitrogen). Luciferase activity was measured using Dual‐Luciferase Reporter assays (E1910, Promega, USA). Briefly, trophoblast cells were incubated for 24 h and lyzed in passive lysis buffer on ice for 20 min. Cell lysates were then mixed with luciferase assay reagent. The firefly luciferase activity was measured on a fluorescence analyzer (PE envision, PE, China). Subsequently, cell lysates were incubated with stop reagent and the renilla luciferases activity was measured as background value.
Pmirglo basic reporter vector
Manufactured by Promega
Sourced in United States
The PmirGLO‐basic reporter vector is a plasmid designed for gene expression studies. It contains a multiple cloning site for inserting a gene of interest, along with a luciferase reporter gene for monitoring transcriptional activity. The vector is suitable for a variety of cell lines and applications requiring a basic reporter system.
Automatically generated - may contain errors
2 protocols using pmirglo basic reporter vector
1
Investigating BRCA1 and lnc-HZ10 Promoters
2
Identifying miR-150-5p Binding to PVT1
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StarBase v2.0 database (41 (link)) was used to analyze the potential miRNAs that interact with PVT1. miR-150-5p was identified as a potential miRNA. Human PVT1-3′-untranslated region (3′-UTR) and GLUT-1-3′-UTR reporter constructs containing the potential binding site of miR-150-5p, and their identical sequences with a mutation in the seed sequence, which were constructed into pmiR-GLO-basic reporter vector (Promega Corp.). Cells (2×105 cells/well) were plated into 48-well plates for 12 h, co-transfected with the indicated plasmids and miR-150-5p mimics for 24 h. The plasmids (200 ng) were mixed with Lipofectamine 2000 (Invitrogen; Thermo Fisher Scientific, Inc.), Renilla luciferase plasmids and DMEM for 30 min and then added into the prepared cells for 24 h. Finally, the luciferase activities were measured by the Dual-Luciferase Assay kit (Promega Corp.) according to the manufacturer's instructions. Every sample was tested for three times and the whole experiment was repeated for three times.
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