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Plasmid Files

pCBG99-Control

Control vector with the SV40 enhancer and promoter driving strong expression of click beetle green luciferase.

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pCBG99-Control Sequence and MappCBG99-Control.dna
Map and Sequence File   
Sequence Author:  Promega
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 BsgI (5222) AarI - BfuAI - BspMI (5194) RVprimer3 (5173 .. 5192) poly(A) signal NotI (5064) BsaAI (4715) XmnI (4165) AseI (3738) AhdI (3566) AlwNI (3089) PciI (2673) Acc65I (1) KpnI (5) Eco53kI (9) SacI (11) NheI (21) BmtI (25) TspMI - XmaI (26) SmaI - SrfI (28) PaeR7I - XhoI (32) BglII (36) SfiI (182) StuI (228) AvrII (229) HindIII (245) BtgI - NcoI (278) PflMI (330) BstXI (484) ZraI (504) AatII (506) PfoI * (683) PspOMI (1416) ApaI (1420) EcoO109I (1450) XbaI (1909) FseI (1928) HpaI (2069) SexAI * (2319) BamHI (2417) SalI (2423) PshAI (2488) RVprimer4 (2474 .. 2493) BspQI - SapI (2557) pCBG99-Control 5231 bp
BsgI  (5222)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14

Efficient cleavage requires at least two copies of the BsgI
recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
AarI  (5194)
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 AarI
recognition sequence.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its
electrophoretic mobility.
BfuAI  (5194)
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  (5194)
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.
NotI  (5064)
1 site
G C G G C C G C C G C C G G C G
BsaAI  (4715)
1 site
Y A C G T R R T G C A Y
XmnI  (4165)
1 site
G A A N N N N T T C C T T N N N N A A G
AseI  (3738)
1 site
A T T A A T T A A T T A
AhdI  (3566)
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.
AlwNI  (3089)
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.
PciI  (2673)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
Acc65I  (1)
1 site
G G T A C C C C A T G G
KpnI  (5)
1 site
G G T A C C C C A T G G
Eco53kI  (9)
1 site
G A G C T C C T C G A G
SacI  (11)
1 site
G A G C T C C T C G A G
NheI  (21)
1 site
G C T A G C C G A T C G
BmtI  (25)
1 site
G C T A G C C G A T C G
TspMI  (26)
1 site
C C C G G G G G G C C C
XmaI  (26)
1 site
C C C G G G G G G C C C

Efficient cleavage requires at least two copies of the XmaI
recognition sequence.
Full cleavage with XmaI may require a long incubation.
SmaI  (28)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
SrfI  (28)
1 site
G C C C G G G C C G G G C C C G
PaeR7I  (32)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (32)
1 site
C T C G A G G A G C T C
BglII  (36)
1 site
A G A T C T T C T A G A
SfiI  (182)
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  (228)
1 site
A G G C C T T C C G G A
AvrII  (229)
1 site
C C T A G G G G A T C C
HindIII  (245)
1 site
A A G C T T T T C G A A
BtgI  (278)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
NcoI  (278)
1 site
C C A T G G G G T A C C
PflMI  (330)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
BstXI  (484)
1 site
C C A N N N N N N T G G G G T N N N N N N A C C

Sticky ends from different BstXI sites may not be compatible.
ZraI  (504)
1 site
G A C G T C C T G C A G
AatII  (506)
1 site
G A C G T C C T G C A G
PfoI  (683)
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.
PspOMI  (1416)
1 site
G G G C C C C C C G G G
ApaI  (1420)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
EcoO109I  (1450)
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  (1909)
1 site
T C T A G A A G A T C T
FseI  (1928)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
HpaI  (2069)
1 site
G T T A A C C A A T T G
SexAI  (2319)
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.
BamHI  (2417)
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.
SalI  (2423)
1 site
G T C G A C C A G C T G
PshAI  (2488)
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.
BspQI  (2557)
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  (2557)
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.
RVprimer3
20-mer  /  50% GC
1 binding site
5173 .. 5192  =  20 annealed bases
Tm  =  54°C
RVprimer4
20-mer  /  65% GC
1 binding site
2474 .. 2493  =  20 annealed bases
Tm  =  61°C
CBG99luc
280 .. 1908  =  1629 bp
542 amino acids  =  60.1 kDa
Product: green-emitting variant of click beetle
luciferase
human codon-optimized
CBG99luc
280 .. 1908  =  1629 bp
542 amino acids  =  60.1 kDa
Product: green-emitting variant of click beetle
luciferase
human codon-optimized
AmpR
3493 .. 4353  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   3493 .. 4284  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3493 .. 4353  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4285 .. 4353  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3493 .. 4353  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
2734 .. 3322  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2734 .. 3322  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
4485 .. 4940  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
4485 .. 4940  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
SV40 promoter
48 .. 244  =  197 bp
SV40 early promoter
SV40 promoter
48 .. 244  =  197 bp
SV40 early promoter
SV40 poly(A) signal
1948 .. 2069  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1948 .. 2069  =  122 bp
SV40 polyadenylation signal
AmpR promoter
4354 .. 4458  =  105 bp
AmpR promoter
4354 .. 4458  =  105 bp
pause site
5133 .. 5224  =  92 bp
RNA polymerase II transcriptional pause signal from
the human α2 globin gene
pause site
5133 .. 5224  =  92 bp
RNA polymerase II transcriptional pause signal from
the human α2 globin gene
poly(A) signal
5071 .. 5119  =  49 bp
synthetic polyadenylation signal
poly(A) signal
5071 .. 5119  =  49 bp
synthetic polyadenylation signal
SV40 ori
95 .. 230  =  136 bp
SV40 origin of replication
SV40 ori
95 .. 230  =  136 bp
SV40 origin of replication
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