pGL4 Vectors are state-of-the-art Luciferase Reporter Vectors. This vector family features codon optimized synthetic genes with or without Rapid Response™ technology, and clean backbones with dramatically reduced numbers of cryptic regulatory elements. Together, these features provide superior sensitivity, reliability and biological relevance. Combine a pGL4 Vector with any of the Promega Luciferase Assay Systems such as EnduRen™, VivRen™, Dual-Glo™, Bright-Glo™ or Steady-Glo® to create an ultra-sensitive cell-based assay that enables you to accurately probe the regulation and activity of a protein or a pathway inside the cell or in a cell lysate.
The pGL4 Vector Family includes:
Figure 1. The pGL4 Luciferase Reporter Vector Family offers a
common configuration with multiple options to match your needs.
The pGL4 vectors multiple cloning region (MCR) is based on the MCR
in our pGL3 Vectors. The two Sfi I restriction sites enable easy
transfer of DNA between the pGL4 Vectors.
Figure2. New synthetic firefly luciferase luc2 gene displays
higher expression and has fewer transcription factor binding sites
than the luc+ gene.
A. Comparison of relative expression between optimized synthetic
firefly gene (luc2) and the previous version (luc+).
The optimized synthetic firefly gene (luc2) was cloned into
the pGL3-Control Vector, replacing the luc+ gene. Each vector
was separately transfected along with a phRL-TK control into NIH/3T3,
CHO, HEK 293 and HeLa cells. Twenty-four hours later, cells were
lysed and luminescence was measured using the Dual-Luciferase®
Reporter Assay System. Relative light units were normalized to Renilla
luciferase expression from the phRL-TK control.
B. luc2 has approximately 90% fewer transcription factor
binding sites than luc+. A comparison of the known consensus
sequence transcription factor binding sites in the synthetic firefly
luc2 and the luc+ genes are presented.
Figure3. Synthetic Renilla luciferase hRluc gene
exhibits higher expression and has fewer transcription factor binding
sites than the native gene.
A. Comparison of relative expression between optimized synthetic
Renilla gene (hRluc) and the previous native version
(Rluc). To compare relative expression, CHO and HeLa
cells were transfeted with pGL3-Control Vector (with SV40 promoter
and enhancer) encoding either the synthetic (hRluc) or the
native gene (Rluc). Twenty-four hours later, cells were lysed
and Renilla luciferase measured using the Dual-Luciferase®
Reporter Assay System.
B. hRluc has approximately 95% fewer sites than Rluc.
A comparison of the known consensus sequence transcription factor
binding sites in the synthetic Renilla luciferase (hRluc)
and the native (Rluc) genes are presented.
Figure 4. Comparison of pGL3 and pGL4 Vector Backbones. pGL3
and pGL4 Vector backbones are shown with known transcriptional binding
sites upstream from the reporter gene coding region. The number
of sites was reduced from over 200 in the pGL3 vectors to less than
50 in the pGL4 vectors. While still maintaining functionality, 6
promoter modules/potential transcription start sites were removed
from the pGL4 vector backbone.
A. pGL4 Rapid Response™ Reporter Gene Design. pGL4 Rapid
Response™ Vectors incorporate fusions of protein degradation signals
hPEST and hCL1 to the C-terminus of luciferase.
B. Destablized luciferases increase the response and reduce the
time to maximum induction. luc2, firefly luciferase;
luc2P, firefly luciferase with PEST sequence; lucCP,
firefly luciferase with hCL1 and hPEST sequences. These luciferases
were used to monitor the response of cAMP Response Element (CRE).
A stable HEK293 cell line with CRE-luc was induced with 1μM
isoproterenol/100μM Ro-20-1724, samples were harvested every hour
to quantify luminescence.
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