pSV40-GLuc

Control vector with a strong constitutive SV40 promoter for expressing secreted Gaussia luciferase in mammalian cells.

Sequence Author: New England Biolabs

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NheI (18) PaqCI (4141) DraIII (3665) BsaAI (3662) BsaHI (3050) AseI (2685) BsaI (2574) BmtI (22) PstI (27) MCS EcoRI (34) EcoRV (42) SfiI (185) BseRI (228) StuI (231) AvrII (232) MscI (399) NruI (411) SacII (437) PspXI (871) XbaI (883) HpaI (1019) MfeI (1028) CsiI - SexAI * (1270) PfoI * (1360) PshAI (1435) AfeI (1496) BspQI - SapI (1504) AflIII - PciI (1620) pSV40-GLuc 4178 bp
NheI  (18)
1 site
G C T A G C C G A T C G
PaqCI  (4141)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the PaqCI recognition sequence.
Sticky ends from different PaqCI sites may not be compatible.
Cleavage can be improved with PaqCI Activator.
DraIII  (3665)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
BsaAI  (3662)
1 site
Y A C G T R R T G C A Y
BsaHI  (3050)
1 site
G R C G Y C C Y G C R G

BsaHI is typically used at 37°C, but is even more active at 60°C.
AseI  (2685)
1 site
A T T A A T T A A T T A
BsaI  (2574)
1 site
G G T C T C N C C A G A G N ( N ) 4

Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
BmtI  (22)
1 site
G C T A G C C G A T C G
PstI  (27)
1 site
C T G C A G G A C G T C
EcoRI  (34)
1 site
G A A T T C C T T A A G
EcoRV  (42)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to delete a base after cleavage.
SfiI  (185)
1 site
G G C C N N N N N G G C C C C G G N N N N N C C G G

Efficient cleavage requires at least two copies of the SfiI recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
BseRI  (228)
1 site
G A G G A G ( N ) 8 N N C T C C T C ( N ) 8

Sticky ends from different BseRI sites may not be compatible.
BseRI quickly loses activity at 37°C.
Prolonged incubation with BseRI may lead to degradation of the DNA.
StuI  (231)
1 site
A G G C C T T C C G G A
AvrII  (232)
1 site
C C T A G G G G A T C C
MscI  (399)
1 site
T G G C C A A C C G G T
NruI  (411)
1 site
T C G C G A A G C G C T
SacII  (437)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII recognition sequence.
PspXI  (871)
1 site
V C T C G A G B B G A G C T C V
XbaI  (883)
1 site
T C T A G A A G A T C T
HpaI  (1019)
1 site
G T T A A C C A A T T G
MfeI  (1028)
1 site
C A A T T G G T T A A C
CsiI  (1270)
1 site
A C C W G G T T G G W C C A

Sticky ends from different CsiI sites may not be compatible.
SexAI  (1270)
1 site
A C C W G G T T G G W C C A
* Blocked by Dcm methylation.
Sticky ends from different SexAI sites may not be compatible.
PfoI  (1360)
1 site
T C C N G G A A G G N C C T
* Blocked by Dcm methylation.
Sticky ends from different PfoI sites may not be compatible.
PshAI  (1435)
1 site
G A C N N N N G T C C T G N N N N C A G

PshAI quickly loses activity at 37°C, but can be used at 25°C for long incubations.
AfeI  (1496)
1 site
A G C G C T T C G C G A
BspQI  (1504)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (1504)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be mixed before removing an aliquot.
AflIII  (1620)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
PciI  (1620)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AmpR
2440 .. 3300  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   2440 .. 3231  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2440 .. 3300  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   3232 .. 3300  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2440 .. 3300  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
1681 .. 2269  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
1681 .. 2269  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
hGLuc
306 .. 863  =  558 bp
185 amino acids  =  19.9 kDa
Product: secreted Gaussia luciferase
human codon-optimized
hGLuc
306 .. 863  =  558 bp
185 amino acids  =  19.9 kDa
Product: secreted Gaussia luciferase
human codon-optimized
f1 ori
3432 .. 3887  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
3432 .. 3887  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
SV40 promoter
52 .. 246  =  195 bp
SV40 early promoter
SV40 promoter
52 .. 246  =  195 bp
SV40 early promoter
SV40 poly(A) signal
898 .. 1019  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
898 .. 1019  =  122 bp
SV40 polyadenylation signal
AmpR promoter
3301 .. 3405  =  105 bp
AmpR promoter
3301 .. 3405  =  105 bp
pause site
4080 .. 4171  =  92 bp
RNA polymerase II transcriptional pause signal from the human α2 globin gene
pause site
4080 .. 4171  =  92 bp
RNA polymerase II transcriptional pause signal from the human α2 globin gene
poly(A) signal
4018 .. 4066  =  49 bp
synthetic polyadenylation signal
poly(A) signal
4018 .. 4066  =  49 bp
synthetic polyadenylation signal
MCS
18 .. 45  =  28 bp
multiple cloning site
MCS
18 .. 45  =  28 bp
multiple cloning site
SV40 ori
98 .. 233  =  136 bp
SV40 origin of replication
SV40 ori
98 .. 233  =  136 bp
SV40 origin of replication
ORF:  2570 .. 2836  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  306 .. 863  =  558 bp
ORF:  185 amino acids  =  19.9 kDa
ORF:  984 .. 1469  =  486 bp
ORF:  161 amino acids  =  18.0 kDa
ORF:  233 .. 574  =  342 bp
ORF:  113 amino acids  =  12.0 kDa
ORF:  2440 .. 3300  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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