phRL-CMV

Mammalian vector for strong constitutive expression of humanized Renilla luciferase.

Sequence Author: Promega

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HaeII (3828) PspFI (3768) BseYI (3764) AlwNI (3659) AhdI (3180) BsrFI (3095) FspI (2957) PvuI (2811) BamHI (2257) BspDI - ClaI (2250) MfeI (2164) HpaI (2155) PsiI (2135) ApoI (2100) EagI - NotI (2012) BglII (1) BsrGI (102) SpeI (158) NdeI (393) BtgZI (493) BsaAI - SnaBI (499) Eco53kI (725) SacI (727) HindIII (754) PstI (836) BfuAI - BspMI (850) BbsI (934) NheI (1058) BmtI (1062) AvaI - BsoBI (1089) DraIII (1256) NsiI (1287) PfoI * (1348) EcoRV (1553) Bsu36I (1654) NruI (1729) PflFI - Tth111I (1764) PasI (1842) EcoO109I (1913) XbaI (2005) phRL-CMV 4079 bp
HaeII  (3828)
1 site
R G C G C Y Y C G C G R
PspFI  (3768)
1 site
C C C A G C G G G T C G
BseYI  (3764)
1 site
C C C A G C G G G T C G

After cleavage, BseYI can remain bound to DNA and alter its electrophoretic mobility.
AlwNI  (3659)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
AhdI  (3180)
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.
BsrFI  (3095)
1 site
R C C G G Y Y G G C C R

Cleavage may be enhanced when more than one copy of the BsrFI recognition sequence is present.
After cleavage, BsrFI can remain bound to DNA and alter its electrophoretic mobility.
FspI  (2957)
1 site
T G C G C A A C G C G T
PvuI  (2811)
1 site
C G A T C G G C T A G C
BamHI  (2257)
1 site
G G A T C C C C T A G G

After cleavage, BamHI-HF® (but not the original BamHI) can remain bound to DNA and alter its electrophoretic mobility.
BspDI  (2250)
1 site
A T C G A T T A G C T A
ClaI  (2250)
1 site
A T C G A T T A G C T A
MfeI  (2164)
1 site
C A A T T G G T T A A C
HpaI  (2155)
1 site
G T T A A C C A A T T G
PsiI  (2135)
1 site
T T A T A A A A T A T T
ApoI  (2100)
1 site
R A A T T Y Y T T A A R

ApoI is typically used at 50°C, but is 50% active at 37°C.
EagI  (2012)
1 site
C G G C C G G C C G G C
NotI  (2012)
1 site
G C G G C C G C C G C C G G C G
BglII  (1)
1 site
A G A T C T T C T A G A
BsrGI  (102)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
SpeI  (158)
1 site
A C T A G T T G A T C A
NdeI  (393)
1 site
C A T A T G G T A T A C

Prolonged incubation with NdeI may lead to removal of additional nucleotides.
BtgZI  (493)
1 site
G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4

Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
BsaAI  (499)
1 site
Y A C G T R R T G C A Y
SnaBI  (499)
1 site
T A C G T A A T G C A T
Eco53kI  (725)
1 site
G A G C T C C T C G A G
SacI  (727)
1 site
G A G C T C C T C G A G
HindIII  (754)
1 site
A A G C T T T T C G A A
PstI  (836)
1 site
C T G C A G G A C G T C
BfuAI  (850)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BfuAI recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (850)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BspMI recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
BbsI  (934)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
NheI  (1058)
1 site
G C T A G C C G A T C G
BmtI  (1062)
1 site
G C T A G C C G A T C G
AvaI  (1089)
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  (1089)
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.
DraIII  (1256)
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.
NsiI  (1287)
1 site
A T G C A T T A C G T A
PfoI  (1348)
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.
EcoRV  (1553)
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.
Bsu36I  (1654)
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.
NruI  (1729)
1 site
T C G C G A A G C G C T
PflFI  (1764)
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  (1764)
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.
PasI  (1842)
1 site
C C C W G G G G G G W C C C

Sticky ends from different PasI sites may not be compatible.
EcoO109I  (1913)
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.
XbaI  (2005)
1 site
T C T A G A A G A T C T
hRluc
1068 .. 2003  =  936 bp
311 amino acids  =  36.0 kDa
Product: Renilla luciferase
human codon-optimized
hRluc
1068 .. 2003  =  936 bp
311 amino acids  =  36.0 kDa
Product: Renilla luciferase
human codon-optimized
AmpR
2393 .. 3253  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   2393 .. 2461  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2393 .. 3253  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   2462 .. 3253  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2393 .. 3253  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
3424 .. 4012  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3424 .. 4012  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
CMV enhancer
220 .. 523  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
220 .. 523  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
524 .. 727  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
524 .. 727  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
chimeric intron
863 .. 995  =  133 bp
chimera between introns from human β-globin and immunoglobulin heavy chain genes
chimeric intron
863 .. 995  =  133 bp
chimera between introns from human β-globin and immunoglobulin heavy chain genes
SV40 poly(A) signal
2034 .. 2155  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2034 .. 2155  =  122 bp
SV40 polyadenylation signal
AmpR promoter
2288 .. 2392  =  105 bp
AmpR promoter
2288 .. 2392  =  105 bp
T7 promoter
1040 .. 1058  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
1040 .. 1058  =  19 bp
promoter for bacteriophage T7 RNA polymerase
ORF:  2393 .. 3253  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  1068 .. 2003  =  936 bp
ORF:  311 amino acids  =  36.0 kDa
ORF:  2857 .. 3123  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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