2012 Promega catalogue
2012 Promega catalogue
2012 Promega catalogue
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Life<br />
Science<br />
Catalog<br />
<strong>2012</strong><br />
Worldwide Contact List<br />
Section<br />
Contents<br />
Table of<br />
Contents<br />
Cell Signaling<br />
140<br />
GloResponse Luciferase Reporter Cell<br />
Lines<br />
Product Size Cat.# Price ($)<br />
GloResponse CRE-luc2P HEK293 Cell Line 2 vials E8500 Pls. Enq.<br />
GloResponse NFAT-RE-luc2P HEK293 Cell Line 2 vials E8510 Pls. Enq.<br />
GloResponse NF-κB-RE-luc2P HEK293 Cell Line 2 vials E8520 Pls. Enq.<br />
GloResponse 9XGAL4UAS-luc2P HEK293 Cell Line 2 vials E8530 Pls. Enq.<br />
For Research Use Only. Not for Use in Diagnostic Procedures.<br />
Description: The GloResponse Luciferase Reporter Cell Lines contain<br />
optimized, state-of-the-art luciferase reporter technology integrated into a<br />
cell line. This allows the rapid development of a reporter assay based on the<br />
pathway of interest regulating the luciferase gene. Assays configured using the<br />
GloResponse Cell Lines are amenable for high-throughput screening. These<br />
assays typically have greater response dynamics (fold of induction) than other<br />
assay formats and good quality as indicated by the high Z´ values.<br />
GloResponse Cell Lines were developed to study a variety of signaling<br />
pathways. Activators of these pathways may be native to the HEK293 cell line.<br />
Activity of non-native activators can be studied after they have been introduced<br />
by transfection.<br />
GPCRs regulate a wide-range of biological functions and are one of the most<br />
important target classes for drug discovery. GPCR signaling pathways can be<br />
categorized into three classes based on the G protein α-subunit involved: Gs,<br />
Gi/o and Gq. The GloResponse CRE-luc2P HEK293 Cell Line can be used to<br />
study and configure screening assays for Gs- and Gi/o-coupled GPCRs, which<br />
signal through cAMP and the cAMP Response Element (CRE). For Gq-coupled<br />
GPCRs, which signal through calcium ion release and activate the Nuclear<br />
Factor of Activated T-Cells response element (NFAT-RE), the GloResponse<br />
NFAT-RE-luc2P HEK293 Cell Line should be used.<br />
NF-κB-REs are the DNA binding sequences for the NF-κB transcription factor<br />
complex, which is responsible for regulating inflammation, immune response,<br />
cell growth and apoptosis. The GloResponse NF-κB-RE-luc2P HEK293 Cell<br />
Line is designed for rapid and convenient analysis of any cellular response that<br />
results in modulation of NF-κB activities.<br />
The GloResponse 9XGAL4UAS-luc2P HEK293 Cell Line contains nine<br />
repeats of GAL4 UAS (Upstream Activator Sequence) driving the transcription<br />
of the luciferase reporter gene luc2P in response to binding of a fusion protein<br />
containing the GAL4 DNA Binding Domain, such as the Estrogen Receptor<br />
Ligand Binding Domain in pBIND-ERα Vector (Cat.# E1390) when activated by<br />
a ligand. This makes the cell line suitable for the study of nuclear receptors or<br />
can be used to study other types of protein:protein and protein:DNA<br />
interactions. The GAL4 DNA Binding Domain partner must be introduced to this<br />
cell line by transfection or other similar techniques.<br />
The GloResponse Cell Lines were generated by clonal selection of HEK293<br />
cells stably transfected with pGL4-based vectors carrying specific response<br />
elements for the pathway of interest. These cell lines incorporate the<br />
improvements developed for the pGL4 family of reporter vectors for enhanced<br />
performance. The destabilized luc2P luciferase reporter is used for improved<br />
responsiveness to transcriptional dynamics. The luc2P gene is codon optimized<br />
for enhanced expression in mammalian cells, and the pGL4 plasmid backbone<br />
was engineered to reduce background reporter expression. The result is a cell<br />
line with very high induction levels when the pathway of interest is activated.<br />
Features:<br />
• Convenient: Prebuilt, optimized luciferase reporter cell lines.<br />
• Robust: Large assay window provided by high levels of induction and low<br />
background expression.<br />
• Faster Results: Improved responsiveness to transcriptional dynamics with<br />
destabilized luciferase.<br />
Storage Conditions: Place frozen cells in storage at less than or equal to<br />
–140°C (mechanical deep freeze or vapor phase liquid nitrogen) until you are<br />
ready to thaw and propagate them. We strongly recommend that the cells are<br />
propagated, using the provided procedure, as soon as possible. This will ensure<br />
the optimal cell viability and assay performance.<br />
Protocol Part#<br />
GloResponse CRE-luc2P HEK293 Cell Line Technical Bulletin TB362<br />
Protocol Part#<br />
GloResponse NFAT-RE-luc2P HEK293 Cell Line Technical Bulletin TB363<br />
GloResponse NF-κB-RE-luc2P HEK293 Cell Line Technical<br />
Bulletin<br />
TB380<br />
GloResponse 9XGAL4UAS-luc2P HEK293 Cell Line Technical<br />
Bulletin<br />
TB552<br />
pGL4-RE-luc2P<br />
RE luc2P PSV40 Hyg r<br />
pRluc-Neo r-GPCR<br />
PCMV GPCR PSV40 Rluc-Neo r<br />
Two plasmids involved in the dual-luciferase GPCR assay. RE, response<br />
element/promoter; luc2P, destabilized firefly luciferase with PEST sequence;<br />
P SV40, SV40 promoter; Hyg r , hygromycin resistance gene; P CMV, CMV promoter;<br />
Rluc-neo r , Renilla luciferase and neomycin resistance gene fusion. PEST<br />
sequences are associated with rapidly degraded proteins.<br />
Luminescence (RLU)<br />
3 × 10 5<br />
2 × 10 5<br />
1 × 10 5<br />
pergolide<br />
apomorphine<br />
SKF38393<br />
dopamine<br />
dihydrexidine<br />
0<br />
–9 –8 –7<br />
log [agonist], M<br />
–6 –5<br />
Ranking compound potency and detection of DRD1 partial agonists. A<br />
GloResponse CRE-luc2P clone stably expressing dopamine receptor D1 was<br />
plated at 10,000 cells/well in a 96-well plate. Each agonist was serially diluted<br />
1:2, then added to wells in replicates of four, beginning with 50μM. Cells<br />
were incubated with agonist for four hours, harvested and analyzed using the<br />
Dual-Glo Luciferase Assay System (Cat.# E2920). Luciferase activity was<br />
measured on the GloMax ® 96 Microplate Luminometer (Cat.# E6501).<br />
Custom Assay Services<br />
Biology-driven, <strong>Promega</strong> technology-enabled<br />
custom solutions for :<br />
• Cell Engineering<br />
• Assay Development & Qualification<br />
• Assay-Ready Cells In-Scale<br />
www.promega.com/CAS/<br />
For complete and up-to-date product information visit: www.promega.com/catalog<br />
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