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pGreenFire Lenti-Reporter BamHI mCMV Spe I Xba I EcoRI Cla I cPPT 3’ΔLTR WPRE SV40 ORI pUC ori Amp R Nde I RSV 5’LTR Ψ SV40 Poly-A copGFP Luciferase T2A peptide Multiple Cloning Site for TRE Monitor Pathway Activation with Transcriptional Reporters SBI’s lentiviral-based reporter system is a novel approach to study transcriptional regulation and offers many advantages over current transcription reporter systems. pGreenFire1 (pGF1) is a versatile HIV- based transcription reporter that co-expresses destabilized copGFP and Firefly Luciferase enabling the simultaneous detection of GFP and Luciferase signals for quantitative transcription activation response. The activation of a signal transduction pathway (e.g. by growth factors, drugs, etc.) can be monitored by the expression level of the reporter gene controlled by a promoter containing the corresponding signal response elements. Copy number of reporter constructs can be controlled by varying the multiplicity of infection (MOI). Commonly used plasmid-based transcriptional reporter vectors often skew transcriptional network reporting due to their episomal nature. SBI’s lentivector-based transcription reporters integrate into the host’s genome and enable proper chromatinization to produce more faithful transcriptional activity reporting. Highlights • Dual reporter vector system to quantitate Firefly Luciferase and GFP for live cell imaging • Ready-to-use pre-packaged constructs with a wide range of Transcriptional Response Elements (TREs) • Study transactivation and epigenetic effects more accurately • Low background with robust cell transcription activation signals • Establish stable reporter cell lines pGreenFire1 (pGF1) is a versatile HIV-based lentivector that co-expresses destabilized copGFP and Firefly Luciferase enabling the detection of both GFP signals as well as Luciferase for quantitative transcription activation reporter assays. Promoter is active when the specific Transcription Factor is bound to the TRE sequences Minimal CMV promoter is inactive without activation Lentiviral Technology www.systembio.com/greenfire Accurately monitor and quantitate transcription network activity mCMV TRE-specifc Activator mCMV GFP Reporter TRE TRE TRE TRE Binding of Specific Transcription Factors to TRE sequences TRE-specifc Activator TRE-specifc Activator TRE-specifc Activator TRE TRE TRE TRE Luciferase Reporter Luciferase Reporter Transcription Factor Response Elements Transcription Factor Response Elements GFP Reporter pGreenFire Transcription Reporters Lentiviral Transcriptional Reporter Systems
2

pGreenFire™ Transcription Reporters

Feb 14, 2017

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Page 1: pGreenFire™ Transcription Reporters

pGreenFireLenti-Reporter

BamHI

mCMVSpe I

Xba IEcoRI

Cla IcPPT

3’ΔLTRWPRE

SV40 ORI

pUC ori

AmpR

Nde I RSV 5’LTRΨ

SV40 Poly-AcopGFP

LuciferaseT2A peptide

MultipleCloning Site

for TRE

Monitor Pathway Activation with Transcriptional Reporters

SBI’s lentiviral-based reporter system is a novel approach to study transcriptional regulation and offers

many advantages over current transcription reporter systems. pGreenFire1 (pGF1) is a versatile HIV-

based transcription reporter that co-expresses destabilized copGFP and Firefly Luciferase enabling the

simultaneous detection of GFP and Luciferase signals for quantitative transcription activation response.

The activation of a signal transduction pathway (e.g. by growth factors, drugs, etc.) can be monitored by

the expression level of the reporter gene controlled by a promoter containing the corresponding signal

response elements. Copy number of reporter constructs can be controlled by varying the multiplicity of

infection (MOI).

Commonly used plasmid-based transcriptional reporter vectors often skew transcriptional network reporting

due to their episomal nature. SBI’s lentivector-based transcription reporters integrate into the host’s genome

and enable proper chromatinization to produce more faithful transcriptional activity reporting.

Highlights

• Dual reporter vector system to quantitate Firefly Luciferase and GFP for live cell imaging

• Ready-to-use pre-packaged constructs with a wide range of Transcriptional Response Elements (TREs)

• Study transactivation and epigenetic effects more accurately

• Low background with robust cell transcription activation signals

• Establish stable reporter cell lines

pGreenFire1 (pGF1) is a versatile HIV-based lentivector that co-expresses destabilized copGFP and Firefly Luciferase enabling the detection of both GFP signals as well as Luciferase for quantitative transcription activation reporter assays.

Promoter is active when the specific Transcription Factor is bound to the TRE sequences

Minimal CMV promoter is inactive without activation

Lentiviral Technology

www.systembio.com/greenfire

Accurately monitor and quantitate transcription network activity

mCMV

TRE-specifcActivator

mCMV GFP ReporterTRETRETRETRE

Binding of Speci�c Transcription Factors to TRE sequences

TRE-specifcActivator

TRE-specifcActivator

TRE-specifcActivator

TRETRETRETRE Luciferase Reporter

Luciferase Reporter

Transcription Factor Response Elements

Transcription Factor Response Elements

GFP Reporter

pGreenFire™ Transcription ReportersLentiviral Transcriptional Reporter Systems

Page 2: pGreenFire™ Transcription Reporters

Pre-made pGreenFire Transcription Response Element Constructs

None AP1 LXRE HIF1 MEF2 NFAT SMAD

CMV STAT1 Nanog GAS Pax6 CREB Oct 4

NF-κB ISRE Notch SREBP PPARg SP1 C/EBPa

TCF/LEF1 GAL4 p53 SRF cJun RARE

We Also Offer Custom ServicesSystem Biosciences offers a wide-range of custom services to support your research, allowing you to spend less time making tools, and more time making discoveries. To learn more, visit our website at www.systembio.com/service or call us at 888-266-5066.

© 2009 System Biosciences, Inc. All rights reserved. System Biosciences and the System Biosciences logo are trademarks of System Biosciences, Inc.

System Biosciences, Inc.1616 North Shoreline Blvd.Mountain View, CA 94043

Toll Free: 888.266.5066Fax: 650-968-2277Email: [email protected]

pGreenFire™ Transcription Reporters

Luciferase

Luciferase

Luciferase

INF-α (U/mL)

PMA (ng/mL)

TO-901317 (nM)

LXRE

AP1

ISRE

Study Lipid Reponse Pathways

pGreenFire-LXRE construct contains the Liver X receptor transcription factor response elements that can be bound by the LXR family of transciption factors. The pGreenFire-LXRE reporter has a strong response to the LXR agonist TO-901317 compound.

Study Oncogenic Pathways

pGreenFire-AP1 construct responds to the Activator Protein-1 (AP1) transcription factor hetero- or homo-dimeric complexes that comprises members of the proto-oncogene Jun protein family (c-Jun, JunB and JunD) and Fos protein family (c-Fos, Fos B, Fra-1 and Fra-2). SBI’s pGreenFire-AP1 Reporter provides accurate response to this pathway and can be induced by the agonist Phorbol-Myristate-Acetate (PMA).

Study Inflammation Pathways

pGreenFire-ISRE construct contains the Interferon-Stimulated Response Element that responds to Type I interferons (IFN-α and -ß ) that mediate signaling through STAT1 and STAT2 components of the JAK/STAT-signal transduction pathways. Robust transactivation response is observed using Interferon-α as the agonist.

Control

Control

Control

GFP

GFP

GFP

+TO-901317 (5 nM)

+PMA (1000 ng/mL)

+INF-α (1000 U/mL)

RLU

0

150000

200000

250000

300000

50000

100000

0 5

ng/mL PMA

RLU

0

150000

200000

250000

300000

50000

100000

0 1000

U/mL INF-α

RLU

0

150000

200000

250000

300000

50000

100000

0 1000

Available as ready-to-transduce virus or plasmid DNA