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Plasmid Files

pGL3-Promoter

SV40 promoter-containing vector for measuring the activity of enhancer sequences with a luciferase assay.

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pGL3-Promoter Sequence and MappGL3-Promoter.dna
Map and Sequence File   
Sequence Author:  Promega
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 KpnI (5) Acc65I (1) BsgI (5001) RVprimer3 (4952 .. 4971) poly(A) signal NotI (4843) DraIII (4497) BsaAI (4494) XmnI (3944) AseI (3517) BsaI (3406) AhdI (3345) AlwNI (2868) Eco53kI (9) SacI (11) MluI (15) MCS NheI (21) BmtI (25) TspMI - XmaI (26) SmaI - SrfI (28) PaeR7I - XhoI (32) BglII (36) SfiI (182) StuI (228) AvrII (229) HindIII (245) NcoI (278) GLprimer2 (281 .. 303) KasI (312) NarI (313) SfoI (314) PluTI (316) PfoI * (387) BstBI (449) BsrGI (770) BclI * (860) SphI (943) XcmI (1015) EcoO109I - PpuMI (1459) SgrAI (1708) XbaI (1934) FseI (1953) HpaI (2094) BamHI (2196) SalI (2202) AccI (2203) PshAI (2267) RVprimer4 (2253 .. 2272) AfeI (2328) PciI (2452) pGL3-Promoter 5010 bp
KpnI  (5)
1 site
G G T A C C C C A T G G
Acc65I  (1)
1 site
G G T A C C C C A T G G
BsgI  (5001)
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).
NotI  (4843)
1 site
G C G G C C G C C G C C G G C G
DraIII  (4497)
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  (4494)
1 site
Y A C G T R R T G C A Y
XmnI  (3944)
1 site
G A A N N N N T T C C T T N N N N A A G
AseI  (3517)
1 site
A T T A A T T A A T T A
BsaI  (3406)
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  (3345)
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  (2868)
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.
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
MluI  (15)
1 site
A C G C G T T G C G C A
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
NcoI  (278)
1 site
C C A T G G G G T A C C
KasI  (312)
1 site
G G C G C C C C G C G G
NarI  (313)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI
recognition sequence.
SfoI  (314)
1 site
G G C G C C C C G C G G
PluTI  (316)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI
recognition sequence.
PfoI  (387)
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.
BstBI  (449)
1 site
T T C G A A A A G C T T
BsrGI  (770)
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.
BclI  (860)
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.
SphI  (943)
1 site
G C A T G C C G T A C G
XcmI  (1015)
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.
EcoO109I  (1459)
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.
PpuMI  (1459)
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.
SgrAI  (1708)
1 site
C R C C G G Y G G Y G G C C R C

Efficient cleavage requires at least two copies of the SgrAI
recognition sequence.
XbaI  (1934)
1 site
T C T A G A A G A T C T
FseI  (1953)
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  (2094)
1 site
G T T A A C C A A T T G
BamHI  (2196)
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  (2202)
1 site
G T C G A C C A G C T G
AccI  (2203)
1 site
G T M K A C C A K M T G

Efficient cleavage with AccI requires ≥13 bp on each side of the
recognition sequence.
Sticky ends from different AccI sites may not be compatible.
PshAI  (2267)
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  (2328)
1 site
A G C G C T T C G C G A
PciI  (2452)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
RVprimer3
20-mer  /  50% GC
1 binding site
4952 .. 4971  =  20 annealed bases
Tm  =  54°C
GLprimer2
23-mer  /  39% GC
1 binding site
281 .. 303  =  23 annealed bases
Tm  =  57°C
RVprimer4
20-mer  /  65% GC
1 binding site
2253 .. 2272  =  20 annealed bases
Tm  =  61°C
luciferase
280 .. 1932  =  1653 bp
550 amino acids  =  60.6 kDa
Product: firefly luciferase
enhanced luc+ version of the luciferase gene
luciferase
280 .. 1932  =  1653 bp
550 amino acids  =  60.6 kDa
Product: firefly luciferase
enhanced luc+ version of the luciferase gene
AmpR
3272 .. 4132  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   3272 .. 4063  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3272 .. 4132  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4064 .. 4132  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3272 .. 4132  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
2513 .. 3101  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin
of replication
ori
2513 .. 3101  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin
of replication
f1 ori
4264 .. 4719  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
4264 .. 4719  =  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
1973 .. 2094  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1973 .. 2094  =  122 bp
SV40 polyadenylation signal
AmpR promoter
4133 .. 4237  =  105 bp
AmpR promoter
4133 .. 4237  =  105 bp
pause site
4912 .. 5003  =  92 bp
RNA polymerase II transcriptional pause signal from
the human α2 globin gene
pause site
4912 .. 5003  =  92 bp
RNA polymerase II transcriptional pause signal from
the human α2 globin gene
poly(A) signal
4850 .. 4898  =  49 bp
synthetic polyadenylation signal
poly(A) signal
4850 .. 4898  =  49 bp
synthetic polyadenylation signal
MCS
1 .. 41  =  41 bp
multiple cloning site
MCS
1 .. 41  =  41 bp
multiple cloning site
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