Pmir glo luciferase gene

Manufactured by Promega

Sourced in United States

The Pmir-GLO luciferase gene is a laboratory tool used for the expression and detection of luciferase, a bioluminescent protein. It provides a reliable and sensitive method for monitoring gene expression in various cellular systems.

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Lab products found in correlation

Hek293 t cells, by Agilent Technologies (2 mentions) Prl tk, by Takara Bio (2 mentions) Synergy 2 multidetector microplate reader, by Agilent Technologies (1 mentions) Synergy 2 multi detection microplate reader, by Agilent Technologies (1 mentions) Dual luciferase reporter assay system, by Promega (1 mentions)

4 protocols using pmir glo luciferase gene

Luciferase Reporter Assay for mRNA 3'-UTR Binding

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Target gene analysis was performed using the online database of StarBase
biological prediction website (http://starbase.sysu.edu.cn/). The full length of the 3′
untranslated region (3′-UTR) of the FOXP1 mRNA or full-length NEAT1 mRNA was
cloned and amplified, then the PCR product was cloned into the polyclonal loci
downstream of the pmir-GLO luciferase gene (Promega, Madison, WI, USA). Cells
co-transfected were in HEK293 T cells (Shanghai, china), and the luciferase
activity was measured using a Synergy 2 Multi-detector Microplate Reader (BioTek
Instruments, Inc., Winooski, VT, USA). The experiment was independently repeated
three times. Data were normalized for transfection efficiency by dividing
firefly luciferase activity by Renilla luciferase activity.

Pan W., Wu A., Yu H., Yu Q., Zheng B., Yang W., Tian D., Gao Y, & Li P. (2020). NEAT1 Negatively Regulates Cell Proliferation and Migration of Neuroblastoma Cells by miR-183-5p/FOXP1 Via the ERK/AKT Pathway. Cell Transplantation, 29, 0963689720943608.

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Dual-Luciferase Reporter Assay for miRNA Targets

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Target gene analysis was performed using the online database of StarBase biological prediction website (http://starbase.sysu.edu.cn/). The full length of the 3′-untranslated region (3′-UTR) of SIRTI or HCG11 was amplified, and then the PCR product was cloned into the polyclonal loci downstream of the pmir-GLO luciferase gene (Promega, Madison, WI, USA). The plasmids were co-transfected into HEK-293T cells (Shanghai, China), and the luciferase activity was measured using a Synergy 2 Multi-detection Microplate Reader (BioTek Instruments, Inc., USA). The experiment was independently repeated three times. Data were normalized for transfection efficiency by dividing firefly luciferase activity from Renilla luciferase activity.

Li D., Liu Y., Gao W., Han J., Yuan R., Zhang M, & Ge Z. (2020). LncRNA HCG11 Inhibits Adipocyte Differentiation in Human Adipose-Derived Mesenchymal Stem Cells by Sponging miR-204-5p to Upregulate SIRT1. Cell Transplantation, 29, 0963689720968090.

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Validating miR-107 Regulation of Dkk-1 Expression

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The targeting relationship between miR-107 and Dkk-1 was predicted using Targetscan database and verified by dual-luciferase reporter gene assay. The full length of 3′UTR region of Dkk-1 amplified gene (Dkk-1 Wt) and PCR products were cloned into the multiple cloning sites downstream of the pmirGLo Luciferase gene (Promega, Madison, WI). Targetscan database was then used to predict the binding site for miR-107 which includes the 756–763 bases of Dkk-1 3’UTR (seed region). The sequence mutation (Dkk-1 Mut) was generated by mutating the sites of seed region. pRL-TK (TaKaRa, Dalian, China) was used as a reference expression vector for railla luciferase. The miR-107 mimics and NC were co-transfected with luciferase reporter vectors (Dkk-1 Wt and Dkk-1 Mut) into MCF-7 cells respectively. The dual-luciferase activity was detected according to the manufacturer's protocol (Promega Corp, Madison, WI). The experiment was repeated 3 times.

Zhang Z.C., Liu J.X., Shao Z.W., Pu F.F., Wang B.C., Wu Q., Zhang Y.K., Zeng X.L., Guo X.D., Yang S.H, & He T.C. (2017). In vitro effect of microRNA-107 targeting Dkk-1 by regulation of Wnt/β-catenin signaling pathway in osteosarcoma. Medicine, 96(27), e7245.

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TGFBR2 Gene 3'UTR Luciferase Assay

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The whole length of 3′UTR of TGFBR2 gene was amplified. PCR products were cloned into multiple cloning sites on the downstream of pmirGLO luciferase gene (Promega, WI, United States) by using restriction enzyme sites, to construct TGFBR2-wild type (WT) (CCAGCUAUGA CCACAUUGCACUU) group. Binding sites of miR-301b-3p and its target gene were predicted by target gene databases. TGFBR2-mutant (MUT) vector (CCAGCUAUGACCACAUAC GUGAU) was constructed by PCR site-directed mutation method. Renilla luciferase expression vector pRL-TK (TaKaRa, Dalian, China) was taken as the internal reference. HEK-293T cells (3 × 10⁵ per well) were co-transfected with mimic NC/miR-301b-3p mimic and luciferase reporter vectors, respectively. Dual Luciferase activity was tested by using Dual-Luciferase Reporter Assay System (Promega, Madison, WI, United States).

Chen J., Cheng L., Zou W., Wang R., Wang X, & Chen Z. (2021). ADAMTS9-AS1 Constrains Breast Cancer Cell Invasion and Proliferation via Sequestering miR-301b-3p. Frontiers in Cell and Developmental Biology, 9, 719993.

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