LARP4 promoter was sub-cloned into pGL3 reporter vector (Invitrogen) to form pGL3-LARP4 promoter which was co-transfected into SKOV3 or A2780 cells with OE-circ_LARP4 or the empty vector. Circ_LARP4-WT/Mut or LARP4-WT/Mut was sub-cloned into pmirGLO dual-luciferase plasmid (Promega, Madison, WI, USA) to construct pmirGLO-circ_LARP4-WT/Mut or pmirGLO-LARP4-WT/Mut which was co-transfected with miR-513b-5p mimics or NC mimics into SKOV3 or A2780 cells. Luciferase activities were examined through Dual-Luciferase Reporter Assay System (Promega).
Pgl3 reporter vector
Manufactured by Thermo Fisher Scientific
The PGL3 reporter vector is a plasmid used in molecular biology research. It contains a firefly luciferase gene that serves as a reporter for gene expression studies. The vector can be used to measure the activity of regulatory DNA sequences in transfected cells.
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Lab products found in correlation
Pmirglo dual luciferase plasmid, by Promega (1 mentions)
Dual luciferase reporter assay system, by Promega (1 mentions)
Pmirglo, by Promega (1 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
Pgl3 promoter vector, by Promega (1 mentions)
Dual luciferase assay kit, by Promega (1 mentions)
2 protocols using pgl3 reporter vector
1
Circ_LARP4 Promoter Regulation
2
Validating let-7c Regulation of HSP70
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To validate whether let-7c directly targets the HSP70 3′-UTR, we performed a dual-luciferase reporter assay. The 3′-UTR fragments of HSP70 containing the let-7c binding sequences, as well as the mutant 3′-UTR fragments were amplified and sub-cloned into the downstream sites of the firefly luciferase (luc) coding sequence in the pGL3 promoter vector (Promega, USA). Next, 0.8 μg of the recombined pGL3 reporter vector and 50 nM of let-7c mimics and let-7c mimics control miRNA, were co-transfected into HEK-293 cells using Lipofectamine 2000 (Invitrogen). The cells were then harvested 48 h post-transfection and the luciferase activities were measured using a dual luciferase assay kit (Promega) following manufacturer’s instructions.
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