Dual glo luciferase

Manufactured by Promega

Sourced in United States

The Dual-Glo Luciferase is a laboratory instrument used to measure the activity of luciferase enzymes. It provides a sensitive and quantitative analysis of gene expression levels in cellular and cell-free systems. The core function of the Dual-Glo Luciferase is to detect and quantify the luminescent signals produced by the activity of luciferase reporter genes.

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Lab products found in correlation

Veritas luminometer, by Promega (5 mentions) Prism 6, by GraphPad (2 mentions) Pgl4.23 luc2 minp vector, by Promega (2 mentions) Amaxa cell line nucleofactor kit 5, by Lonza (2 mentions) Glomax 20 20 luminometer, by Promega (2 mentions) Lipofectamine 2000 reagent, by Thermo Fisher Scientific (2 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions) C myc, by New England Biolabs (1 mentions) Lipofectamine 2000, by OriGene (1 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions) Q5 site directed mutagenesis kit, by New England Biolabs (1 mentions) Prl cmv, by Promega (1 mentions) Lipofectomine 2000, by Thermo Fisher Scientific (1 mentions) Quikchange lightning site directed mutagenesis, by Agilent Technologies (1 mentions) 17β estradiol, by Steraloids (1 mentions) Prl sv40, by Promega (1 mentions) Forskolin, by MP Biomedicals (1 mentions) Dual glo stop glo, by Promega (1 mentions) Neon transfection system, by Thermo Fisher Scientific (1 mentions) Neon system, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

Dual‐Glo Luciferases Assay System Dual-GloTM Luciferase Assay System Dual-GloTM luciferase assay Dual-glo luciferase reporter system Dual-Glo Luciferase Reporter kit Dual-Glo luciferase reagent Nano-Glo® Dual-Luciferase® Reporter Assay System kit Nano-Glo Dual-Luciferase Reporter (NanoDLR) assay system Dual-Glo Luciferase Assay Dual-Glo Luciferase Assay luciferase kit

24 protocols using dual glo luciferase

Investigating CaSR-mediated signaling in HEK cells

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HEK‐CaSR cells were seeded in 48‐well plates and transiently transfected with 100 ng/mL pBI‐CMV2‐GNA11 WT or mutant proteins, 100 ng/mL luciferase construct (either pGL4‐NFAT or pGL4‐SRE) and 10 ng/mL pRL control vector for 48 hours. Cells were incubated in serum‐free media for 12 hours, followed by treatment of cells for 4 hours with 0.1mM to 10mM CaCl₂. Cells were lysed and assays performed with Dual‐Glo luciferase (Promega, Madison, WI, USA) on a Veritas Luminometer (Promega), as described.19 Luciferase:renilla ratios were expressed as fold‐changes relative to responses at basal CaCl₂ concentrations. For studies with cinacalcet (Cambridge Bioscience, Cambridge, UK), the drug was added to cells, as described.10, 19 All assays were performed using four biological replicates on up to three independent occasions. Statistical analysis was performed by two‐way ANOVA with Tukey's multiple‐comparisons test using GraphPad Prism 6.

Gorvin C.M., Hannan F.M., Cranston T., Valta H., Makitie O., Schalin‐Jantti C, & Thakker R.V. (2017). Cinacalcet Rectifies Hypercalcemia in a Patient With Familial Hypocalciuric Hypercalcemia Type 2 (FHH2) Caused by a Germline Loss‐of‐Function Gα11 Mutation. Journal of Bone and Mineral Research, 33(1), 32-41.

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YAP Activity Measurement in MSCs

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Sub-confluent MSCs on a 6 cm plastic dish were transfected with 8×GTIIC-luciferase (reporter vector of YAP, a gift from Stefano Piccolo, Addgene plasmid # 34615)(24 (link)) and pRL-TK (control vector, Promega, E2241) with Lipofectamine 3000 (Life Technologies, L3000008). After 1 day, the cells were detached and seeded on a 0.5 mL collagen gel prepared in a 12 well culture plate as previously described. Luciferase activity was determined after 3 days in culture with Dual Glo Luciferase (Promega, E2920). Relative activity of YAP was calculated by normalizing the activity of YAP with the activity of HSV-thymidine kinase promoter.

Ishihara S., Inman D.R., Li W.J., Ponik S.M, & Keely P.J. (2017). Mechano-signal transduction in mesenchymal stem cells induces prosaposin secretion to drive the proliferation of breast cancer cells. Cancer research, 77(22), 6179-6189.

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c-MYC Transcriptional Reporter Assay

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HEK293T cells were seeded at 35,000 cells/well in a 96-well plate. Cells were transfected with the MYC luciferase reporter according to protocol described above using Lipofectamine 2000 (Invitrogen, 11668019) as a transfection reagent. Cells were also transfected with either FLAG-eGFP, FLAG-tagged wild-type c-MYC (Origene, RC201611) or C171Y cMYC mammalian expression plasmids using Lipofectamine 2000. Cells were transfected with 1 μg/well of corresponding plasmid in a final ratio of 3:1 Lipofectamine 2000:DNA. The c-MYC C171Y mutant plasmid was generated using Q5 site-directed mutagenesis kit according to the manufacturer’s instructions (New England Biolabs, E0554S). Sequencing was confirmed with Quintara Biosciences. 24 hours post-transfection, media was carefully aspirated from each well, and 50 μL of fresh media containing either DMSO or compound was added. After 24 hours of compound treatment, Dual-Glo luciferase (Promega, E2920) detection was performed according to manufacturer’s protocol. Firefly and Renilla luminescence were read on a SpectraMax i3 plate reader. Background luminescence was subtracted using a blank control then Firefly:Renilla was calculated for each individual well. Lysates were also obtained for confirmation FLAG-protein overexpression by Western blotting.

Boike L., Cioffi A.G., Majewski F.C., Co J., Henning N.J., Jones M.D., Liu G., McKenna J.M., Tallarico J.A., Schirle M, & Nomura D.K. (2020). Discovery of a Functional Covalent Ligand Targeting an Intrinsically Disordered Cysteine Within MYC. Cell chemical biology, 28(1), 4-13.e17.

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Notch Signaling Pathway Assay

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S2 cells in 12-well dishes were transfected when approximately 50% confluent with combinations of pMT-Notch, pMT-DxV5, pMT-Pyd, NRE:Firefly (gift from S. Bray) and Actin:Renilla (gift from G.Merdes). After 24 h, when cells reached 100% confluence, they were re-suspended and seeded into white 96-well plates (Nunc #136101) and CuSO₄ was added after a further 24 h. 24 h after induction, luciferase activity was assayed with Dual-Glo Luciferase (Promega), quantified by a luminometer (Berthold), and Firefly/Renilla ratio was calculated for triplicate samples. For cell-density experiments, cells were re-seeded into white 96-well plates as above (high cell density) or diluted 1/10 with culture medium before reseeding (low cell density). For ligand-induced signalling assays, Notch expressing cells were layered on top of fixed Delta expressing S2 cells (S2-Mt-Dl; DGRC) according to the previously described protocol [4 (link)]. Experiments were repeated a minimum of three times.

Shimizu H., Wilkin M.B., Woodcock S.A., Bonfini A., Hung Y., Mazaleyrat S, & Baron M. (2017). The Drosophila ZO-1 protein Polychaetoid suppresses Deltex-regulated Notch activity to modulate germline stem cell niche formation. Open Biology, 7(4), 160322.

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Calcium-Responsive Transcriptional Activation

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HEK-CaSR cells were seeded in 48-well plates and transiently transfected with 100 ng/ml Gα₁₁ WT or mutant proteins, 100 ng pGL4-SRE luciferase reporter construct, and 10 ng/ml pRL control vector for 48 hours (Promega). Cells were incubated in serum-free media for 12 hours, followed by treatment of cells for 4 hours with 0–10 mM CaCl₂. Cells were lysed and assays performed using Dual-Glo luciferase (Promega) on a Veritas Luminometer (Promega), as previously described (25 (link), 45 (link)).

Gorvin C.M., Hannan F.M., Howles S.A., Babinsky V.N., Piret S.E., Rogers A., Freidin A.J., Stewart M., Paudyal A., Hough T.A., Nesbit M.A., Wells S., Vincent T.L., Brown S.D., Cox R.D, & Thakker R.V. (2017). Gα11 mutation in mice causes hypocalcemia rectifiable by calcilytic therapy. JCI Insight, 2(3), e91103.

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Notch-Responsive Luciferase Assay

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A total of 293 cells were cotransfected transiently with 16 Notch-responsive element CSL (CBF1, Suppressor of Hairless, Lag-1, CSL)/mini-promoter firefly luciferase reporter (constructed by our research team) and constitutively expressed Renilla luciferase reporter (pRL-CMV; Promega, USA). EGCG was added alone or together with ligand-expressing cells (HepG2 cells) 18 h after transfection. Luciferase activities were measured 12–24 h after EGCG addition (Dual Glo Luciferase; Promega). Typically, three replicates were analyzed for each condition, and values were expressed as relative luciferase units (firefly signal divided by the Renilla signal).

Wang T., Xiang Z., Wang Y., Li X., Fang C., Song S., Li C., Yu H., Wang H., Yan L., Hao S., Wang X, & Sheng J. (2017). (−)-Epigallocatechin Gallate Targets Notch to Attenuate the Inflammatory Response in the Immediate Early Stage in Human Macrophages. Frontiers in Immunology, 8, 433.

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Luciferase Assay for siRNA Screening

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RNA interference was performed by using siRNA purchased from Sigma MISSION siRNA library (SI007 siRNA Fluor Universal Negative Control#1, and PDSIRNA2D SASI_Hs01_00150279 siJUN). HEK293T cells were seeded at 35,000 cells/well in a 96-well plate. On Day 0, cells were co-transfected with 150 ng siRNA and myc luciferase reporter per well according to previous protocol using Lipofectomine 2000 (Invitrogen, 11668019). On Day 1, 24 hours post-transfection, 75 μL of fresh media containing vehicle DMSO or EN4 was added to cells in n=6 replicates. After 24 h of compound treatment, Dual-Glo luciferase (Promega, E2920) workup was performed according to manufacturer’s protocol. Firefly and Renilla luminescence were read on a SpectraMax i3 plate reader. Background luminescence was subtracted using a blank control then Firefly:Renilla was calculated for each individual well.

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Ancestral Lophotrochozoan SR LBD Characterization

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The hinge and LBD of the Crassostreas gigas ER were cloned into the pSG5-Gal4DBD vector (a gift of D. Furlow). The resurrected ancestral lophotrochozoan SR LBD sequence was synthesized as a fusion construct containing the hinge domain and C-terminus of the CgER (Genscript, Piscataway, NJ), and cloned into the pSG5-Gal4DBD vector (Fig. S2B). CHO-K1 cells were transfected using Lipofectamine and Plus with 1 ng LBD plasmids, 100 ng of luciferase reporter plasmid (pFRluc) and 0.1 ng of a normalization plasmid (phRLtk). After 4 hours, the transfection mixture was replaced with medium supplemented with stripped serum, and allowed to recover. The cells were treated with 17β-estradiol (Steraloids, Newport, RI) diluted in medium/serum and incubated for 24 hours. Luciferase assays were performed using DualGlo luciferase (Promega, Madison, WI). Mutations were made using QuikChange Lightning Site-directed mutagenesis (Agilent, Englewood, CO), and were verified by sequencing.

Bridgham J.T., Keay J., Ortlund E.A, & Thornton J.W. (2014). Vestigialization of an Allosteric Switch: Genetic and Structural Mechanisms for the Evolution of Constitutive Activity in a Steroid Hormone Receptor. PLoS Genetics, 10(1), e1004058.

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Translocated Enhancer Sequences and Controls

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Five translocated enhancer sequences and five controls with scrambled MYB consensus motifs (replacing CNGTT with GTAAG, see Supplementary Table 6) were synthesized and cloned into the pGL 4.23 [luc2/minP] vector (Promega) by BlueHeron. Enhancer activity was measured in 6 replicates as relative luminescence of the pGL 4.23 [luc2/minP] vector compared to the pGL 4.73 [hRluc/SV40] with Dual-Glo Luciferase (Promega) after a 36 hour co-nucleofection into Jurkat cells following the manufacturer’s instructions (Amaxa cell line nucleofactor kit V from Lonza).

Drier Y., Cotton M.J., Williamson K.E., Gillespie S.M., Ryan R.J., Kluk M.J., Carey C.D., Rodig S.J., Sholl L.M., Afrogheh A.H., Faquin W.C., Queimado L., Qi J., Wick M.J., El-Naggar A.K., Bradner J.E., Moskaluk C.A., Aster J.C., Knoechel B, & Bernstein B.E. (2016). An oncogenic MYB feedback loop drives alternate cell fates in adenoid cystic carcinoma. Nature genetics, 48(3), 265-272.

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Functional Validation of Genetic Variants

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A 1119, bp fragment containing the ABHD8 promoter was cloned into the pGL3 basic luciferase reporter. Reference and risk associated ANKLE1 promoter fragments were synthesized by GenScript and cloned into pGL3 basic. We generated PCR fragments corresponding to PRE A and PRE B and had PRE C haplotype fragments synthesized by GenScript and these were also sub-cloned into ABHD8 and ANKLE1 promoter constructs. PCR primers are listed in Supplementary Table 10. Bre80 and A2780 cells were transiently transfected with equimolar amounts of luciferase reporter constructs using Renilla luciferase as an internal control reporter. Luciferase was measured 24 h after transfection using Dual-Glo Luciferase (Promega). To correct for any differences in transfection efficiency or cell lysate preparation, Firefly luciferase activity was normalized to Renilla luciferase, and the activity of each construct was measured relative to the promoter alone construct, which had a defined activity of 1. Association was assessed by log transforming the data and performing two-way ANOVA, followed by Dunnett's multiple comparisons test; for ease of interpretation, values were back transformed to the original scale for the graphs.

Lawrenson K., Kar S., McCue K., Kuchenbaeker K., Michailidou K., Tyrer J., Beesley J., Ramus S.J., Li Q., Delgado M.K., Lee J.M., Aittomäki K., Andrulis I.L., Anton-Culver H., Arndt V., Arun B.K., Arver B., Bandera E.V., Barile M., Barkardottir R.B., Barrowdale D., Beckmann M.W., Benitez J., Berchuck A., Bisogna M., Bjorge L., Blomqvist C., Blot W., Bogdanova N., Bojesen A., Bojesen S.E., Bolla M.K., Bonanni B., Børresen-Dale A.L., Brauch H., Brennan P., Brenner H., Bruinsma F., Brunet J., Buhari S.A., Burwinkel B., Butzow R., Buys S.S., Cai Q., Caldes T., Campbell I., Canniotto R., Chang-Claude J., Chiquette J., Choi J.Y., Claes K.B., Cook L.S., Cox A., Cramer D.W., Cross S.S., Cybulski C., Czene K., Daly M.B., Damiola F., Dansonka-Mieszkowska A., Darabi H., Dennis J., Devilee P., Diez O., Doherty J.A., Domchek S.M., Dorfling C.M., Dörk T., Dumont M., Ehrencrona H., Ejlertsen B., Ellis S., Engel C., Lee E., Evans D.G., Fasching P.A., Feliubadalo L., Figueroa J., Flesch-Janys D., Fletcher O., Flyger H., Foretova L., Fostira F., Foulkes W.D., Fridley B.L., Friedman E., Frost D., Gambino G., Ganz P.A., Garber J., García-Closas M., Gentry-Maharaj A., Ghoussaini M., Giles G.G., Glasspool R., Godwin A.K., Goldberg M.S., Goldgar D.E., González-Neira A., Goode E.L., Goodman M.T., Greene M.H., Gronwald J., Guénel P., Haiman C.A., Hall P., Hallberg E., Hamann U., Hansen T.V., Harrington P.A., Hartman M., Hassan N., Healey S., Heitz F., Herzog J., Høgdall E., Høgdall C.K., Hogervorst F.B., Hollestelle A., Hopper J.L., Hulick P.J., Huzarski T., Imyanitov E.N., Isaacs C., Ito H., Jakubowska A., Janavicius R., Jensen A., John E.M., Johnson N., Kabisch M., Kang D., Kapuscinski M., Karlan B.Y., Khan S., Kiemeney L.A., Kjaer S.K., Knight J.A., Konstantopoulou I., Kosma V.M., Kristensen V., Kupryjanczyk J., Kwong A., de la Hoya M., Laitman Y., Lambrechts D., Le N., De Leeneer K., Lester J., Levine D.A., Li J., Lindblom A., Long J., Lophatananon A., Loud J.T., Lu K., Lubinski J., Mannermaa A., Manoukian S., Le Marchand L., Margolin S., Marme F., Massuger L.F., Matsuo K., Mazoyer S., McGuffog L., McLean C., McNeish I., Meindl A., Menon U., Mensenkamp A.R., Milne R.L., Montagna M., Moysich K.B., Muir K., Mulligan A.M., Nathanson K.L., Ness R.B., Neuhausen S.L., Nevanlinna H., Nord S., Nussbaum R.L., Odunsi K., Offit K., Olah E., Olopade O.I., Olson J.E., Olswold C., O'Malley D., Orlow I., Orr N., Osorio A., Park S.K., Pearce C.L., Pejovic T., Peterlongo P., Pfeiler G., Phelan C.M., Poole E.M., Pylkäs K., Radice P., Rantala J., Rashid M.U., Rennert G., Rhenius V., Rhiem K., Risch H.A., Rodriguez G., Rossing M.A., Rudolph A., Salvesen H.B., Sangrajrang S., Sawyer E.J., Schildkraut J.M., Schmidt M.K., Schmutzler R.K., Sellers T.A., Seynaeve C., Shah M., Shen C.Y., Shu X.O., Sieh W., Singer C.F., Sinilnikova O.M., Slager S., Song H., Soucy P., Southey M.C., Stenmark-Askmalm M., Stoppa-Lyonnet D., Sutter C., Swerdlow A., Tchatchou S., Teixeira M.R., Teo S.H., Terry K.L., Terry M.B., Thomassen M., Tibiletti M.G., Tihomirova L., Tognazzo S., Toland A.E., Tomlinson I., Torres D., Truong T., Tseng C.C., Tung N., Tworoger S.S., Vachon C., van den Ouweland A.M., van Doorn H.C., van Rensburg E.J., Van't Veer L.J., Vanderstichele A., Vergote I., Vijai J., Wang Q., Wang-Gohrke S., Weitzel J.N., Wentzensen N., Whittemore A.S., Wildiers H., Winqvist R., Wu A.H., Yannoukakos D., Yoon S.Y., Yu J.C., Zheng W., Zheng Y., Khanna K.K., Simard J., Monteiro A.N., French J.D., Couch F.J., Freedman M.L., Easton D.F., Dunning A.M., Pharoah P.D., Edwards S.L., Chenevix-Trench G., Antoniou A.C, & Gayther S.A. (2016). Functional mechanisms underlying pleiotropic risk alleles at the 19p13.1 breast–ovarian cancer susceptibility locus. Nature Communications, 7, 12675.

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