2012 Promega catalogue

Cell Signaling
pSP-luc+NF Fusion Vector
Product Size Cat.# Price ($)
pSP-luc+NF Fusion Vector 20 µg E4471 214.00
For Research Use Only. Not for Use in Diagnostic Procedures.
Description: The pSP-luc+NF Fusion Vector is a luciferase cassette vector
containing the engineered firefly luciferase gene, luc+NF. The luc+NF gene is
related to the luc+ gene found in the pGL3 family of eukaryotic reporter vectors
but has been further modified for maximum flexibility in constructing N-terminal
fusions (NF) with luciferase. Subcloning luc+NF into expression vectors
provides a useful genetic reporter with exceptional sensitivity. The
pSP-luc+NF Fusion Vector is not itself intended for the expression of luciferase
in eukaryotic cells, because it does not contain eukaryotic promoters,
enhancers or polyadenylation signals.
A unique BstEII site has been inserted immediately downstream of the
luciferase ATG translation codon, allowing cloned inserts to be positioned immediately
downstream of the luc+NF initiation codon. This vector is
recommended specifically for applications where N-terminal fusion proteins do
not contain an internal ATG codon at the luciferase junction.
The luc+NF gene is positioned downstream of an SP6 promoter and a
ribosome binding site. An opposing T7 promoter is located immediately
downstream of luc+NF. Thus, the pSP-luc+NF Fusion Vector provides a
convenient template for the in vitro synthesis of both sense and antisense
luciferase transcripts for studies involving in situ hybridization, RNA processing,
RNA transfection or coupled in vitro transcription/translation and protein folding.
Multiple cloning regions containing recognition sequences for commonly used
restriction enzymes are positioned at the 5´ and 3´ ends of luc+NF to provide
maximum flexibility in cloning. Luciferase enzymatic activity can be assayed
most efficiently using one of the Luciferase Assay Systems.
Features:
• Flexibility: Multiple cloning regions are positioned at the 5´ and 3´ ends of
luc to provide maximum flexibility in cloning.
• N-Terminal Fusions with Luciferase: Unique BstEII site located
immediately downstream of the luciferase ATG translation codon.
Storage Conditions: Store at –20°C.
Protocol Part#
pSP-luc+NF Fusion Vector Technical Bulletin TB210
AlwNI 2372
HpaI 1777
1726
1721
1716
1708
1702
T7
XhoI
ClaI
EcoRV
EcoRI
XbaI
1700
luc+ end
➞
Amp r
pSP-luc+NF Fusion
Vector
(4103bp)
BanII 1072
SspI 3653
AatII 3771
NdeI SP6
4020 KpnI
NheI
BglII
AvrII
NarI rbs
81 HindIII
NcoI
Csp45I
217 BstEII
BclI 628
BbuI 711
For complete and up-to-date product information visit: www.promega.com/catalog
➞
15
17
24
30
36
43
45 luc+NF start
47
0752VA08_4A
pSV-β-Galactosidase Control Vector
Product Size Cat.# Price ($)
pSV-β-Galactosidase Control Vector
For Research Use Only. Not for Use in Diagnostic Procedures.
20 µg E1081 167.00
Description: The pSV-β-Galactosidase Control Vector is a positive control
vector for monitoring transfection efficiencies of mammalian cells. The SV40
early promoter and enhancer drive transcription of the lacZ gene, which
encodes the β-galactosidase enzyme. The pSV-β-Galactosidase Control Vector
can be transfected individually or co-transfected with your DNA of interest.
β-galactosidase is an excellent reporter enzyme that can be assayed quickly
and directly in cell extracts using spectrophotometric, fluorescent or
chemiluminescent assays. This reporter enzyme is also widely used for in situ
histochemical analysis using the substrate X-Gal.
The pSV-β-Galactosidase Control Vector can be co-transfected with your DNA
of interest. For example, co-transfection with firefly luciferase gene vectors
(pGL3 Vectors) provide cell extracts that can be assayed for both luciferase
and β-galactosidase activities. In this manner, the pSV-β-Galactosidase Vector
acts as an internal control for transient expression assays. A negative control
extract, prepared from mock-transfected cells, should also be assayed for the
presence of endogenous β-galactosidase activity in cultured cells. In addition,
co-transfection with chloramphenicol acetyltransferase reporter gene vectors
(pCAT ® 3 Vectors) permits assaying for both CAT and β-galactosidase
activities.
The pSV-β-Galactosidase Vector is a modification of pRSV-β-Gal with SV40
and pUC18 sequences substituted for RSV and pBR322 sequences. The
pSV-β-Galactosidase Vector will express β-galactosidase in E. coli due to the
presence of the E. coli gpt promoter located upstream of the lacZ gene.
Colonies of E. coli containing the pSV-β-Galactosidase Vector will appear blue
when plated on media containing X-Gal.
Storage Conditions: Store at –20°C.
Protocol Part#
pSV-ß-Galactosidase Control Vector Technical Bulletin TB094
SV40 Promoter
and Enhancer
EcoRI 6815
HindIII 414
ori
pSV-β-Galactosidase
Vector
(6820bp)
Amp r
lacZ
EcoRI 3701
BamHI 4151
SalI 4163
PstI 4173
0325VA03_3A
277
15
Reporter Assays and Transfection
Section
Contents
Table of
Contents