Dual luciferase reporter assay was performed as previously [13 (link)]. In brief, the 3′-untranslated region (UTR) of CCNG1 (1367BP) mRNA containing the miR-9 binding site were PCR amplified, and cloned into the pmiR-RB-REPORT™ dual luciferase reporter vector (Promega). Site-directed Gene Mutagenesis Kit (Beyotime, Jiangsu, China) was used to produce the mutations of the miR-9 targeting site. The primers and mutation primers were synthesized by RiboBio and listed in Additional file 1 : Table S2. The luciferase activities were measured at 48 h after cotransfection with miRNA mimic or its negtive control (50 nM) and different reporter vectors (50nM). The experiments were performed in triplicate and repeated three times.
Pmir rb report dual luciferase reporter vector
Manufactured by Promega
The PmiR-RB-REPORT™ dual luciferase reporter vector is a tool used for the analysis of microRNA (miRNA) activity. It contains a miRNA response element (MRE) that can be inserted downstream of a reporter gene, enabling the measurement of miRNA-mediated regulation of gene expression.
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Lab products found in correlation
2 protocols using pmir rb report dual luciferase reporter vector
1
Dual Luciferase Assay for miR-9 Targeting
2
Validating miR-138-5p Targeting of LINC01614
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Bioinformatics prediction tools miRDB (http://mirdb.org/ ) and LncBase Predicted determined that miR-138-5p may be a putative target of LINC01614. LINC01614-3′-UTR wild type (WT) and LINC01614-3′-UTR mutant (Mut) were subsequently cloned into pmiR-RB-REPORT dual-luciferase reporter vector (Promega, Madison, Wisconsin). Then cells were cotransfected with miR-138-5p mimic or miR-138-5p mimic negative control and LINC01614-WT or LINC01614-Mut. After 48-hour transfection, proteins were extracted and Dual-Luciferase Reporter Assay Kit was supplied to examine the relative luciferase activities.
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