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Plasmid Files

pSV40-CLuc

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

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pSV40-CLuc Sequence and MappSV40-CLuc.dna
Map and Sequence File   
Sequence Author:  New England Biolabs
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 Acc65I (1) EagI - NotI (5089) DraIII (4743) XmnI (4190) BsaHI (4128) AseI (3763) BsaI (3652) AhdI (3591) KpnI (5) NheI (18) BmtI (22) MCS PstI (27) Eco53kI (30) SacI (32) EcoRI (34) EcoRV (42) HindIII (46) SfiI (185) StuI (231) AvrII (232) AvaI - BsoBI - PaeR7I - XhoI (247) BmeT110I (248) SpeI (272) XcmI (689) BstEII (800) PmlI (1187) BmgBI (1210) EcoO109I - KflI - PpuMI (1299) BclI * (1787) PflFI - Tth111I (1821) Bsu36I (1851) MluI (1953) XbaI (1961) HpaI (2097) MfeI (2106) SalI (2448) AfeI (2574) BspQI - SapI (2582) pSV40-CLuc 5256 bp
Acc65I  (1)
1 site
G G T A C C C C A T G G
EagI  (5089)
1 site
C G G C C G G C C G G C
NotI  (5089)
1 site
G C G G C C G C C G C C G G C G
DraIII  (4743)
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.
XmnI  (4190)
1 site
G A A N N N N T T C C T T N N N N A A G
BsaHI  (4128)
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  (3763)
1 site
A T T A A T T A A T T A
BsaI  (3652)
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.
AhdI  (3591)
1 site
G A C N N N N N G T C C T G N N N N N C A G

The 1-base overhangs produced by AhdI may be hard to ligate.
Sticky ends from different AhdI sites may not be compatible.
KpnI  (5)
1 site
G G T A C C C C A T G G
NheI  (18)
1 site
G C T A G C C G A T C G
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
Eco53kI  (30)
1 site
G A G C T C C T C G A G
SacI  (32)
1 site
G A G C T C C T C G A G
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.
HindIII  (46)
1 site
A A G C T T T T C G A A
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.
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
AvaI  (247)
1 site
C Y C G R G G R G C Y C

Sticky ends from different AvaI sites may not be compatible.
BsoBI  (247)
1 site
C Y C G R G G R G C Y C

Sticky ends from different BsoBI sites may not be compatible.
BsoBI is typically used at 37°C, but can be used at temperatures
up to 65°C.
PaeR7I  (247)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (247)
1 site
C T C G A G G A G C T C
BmeT110I  (248)
1 site
C Y C G R G G R G C Y C
SpeI  (272)
1 site
A C T A G T T G A T C A
XcmI  (689)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
BstEII  (800)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
PmlI  (1187)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
BmgBI  (1210)
1 site
C A C G T C G T G C A G

This recognition sequence is asymmetric, so ligating blunt ends
generated by BmgBI will not always regenerate a BmgBI site.
EcoO109I  (1299)
1 site
R G G N C C Y Y C C N G G R

Sticky ends from different EcoO109I sites may not be compatible.
KflI  (1299)
1 site
G G G W C C C C C C W G G G

Sticky ends from different KflI sites may not be compatible.
PpuMI  (1299)
1 site
R G G W C C Y Y C C W G G R

Sticky ends from different PpuMI sites may not be compatible.
BclI  (1787)
1 site
T G A T C A A C T A G T
* Blocked by Dam methylation.
BclI is typically used at 50-55°C, but is 50% active at 37°C.
PflFI  (1821)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (1821)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
Bsu36I  (1851)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
MluI  (1953)
1 site
A C G C G T T G C G C A
XbaI  (1961)
1 site
T C T A G A A G A T C T
HpaI  (2097)
1 site
G T T A A C C A A T T G
MfeI  (2106)
1 site
C A A T T G G T T A A C
SalI  (2448)
1 site
G T C G A C C A G C T G
AfeI  (2574)
1 site
A G C G C T T C G C G A
BspQI  (2582)
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  (2582)
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.
CLuc
291 .. 1952  =  1662 bp
553 amino acids  =  61.5 kDa
Product: secreted Cypridina luciferase
contains 30 codon substitutions for efficient
translation in mammalian cells
CLuc
291 .. 1952  =  1662 bp
553 amino acids  =  61.5 kDa
Product: secreted Cypridina luciferase
contains 30 codon substitutions for efficient
translation in mammalian cells
AmpR
3518 .. 4378  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   3518 .. 4309  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3518 .. 4378  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4310 .. 4378  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3518 .. 4378  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
2759 .. 3347  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2759 .. 3347  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
4510 .. 4965  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
4510 .. 4965  =  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
1976 .. 2097  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1976 .. 2097  =  122 bp
SV40 polyadenylation signal
AmpR promoter
4379 .. 4483  =  105 bp
AmpR promoter
4379 .. 4483  =  105 bp
pause site
5158 .. 5249  =  92 bp
RNA polymerase II transcriptional pause signal from
the human α2 globin gene
pause site
5158 .. 5249  =  92 bp
RNA polymerase II transcriptional pause signal from
the human α2 globin gene
MCS
1 .. 51  =  51 bp
multiple cloning site
MCS
1 .. 51  =  51 bp
multiple cloning site
poly(A) signal
5096 .. 5144  =  49 bp
synthetic polyadenylation signal
poly(A) signal
5096 .. 5144  =  49 bp
synthetic polyadenylation signal
SV40 ori
98 .. 233  =  136 bp
SV40 origin of replication
SV40 ori
98 .. 233  =  136 bp
SV40 origin of replication
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