pGL4.71[hRlucP]

Promoterless vector encoding destabilized Renilla luciferase for measuring the activity of promoter and enhancer sequences.

Sequence Author: Promega

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BglI - SfiI (8) BsgI (3651) AarI - BfuAI - BspMI (3623) RVprimer3 (3602 .. 3621) BsmBI (3450) SpeI (3437) BstZ17I (3118) SacII (3002) PvuI (2978) MscI (2842) AhdI (2608) BstEII (2533) BstXI - PstI (2530) AleI (2528) NotI (2506) BspHI (2406) AlwNI (2102) BaeGI - Bme1580I (2004) ApaLI (2000) BciVI (1889) BssSI (1859) NspI (1690) Acc65I (14) KpnI (18) Bpu10I (19) Eco53kI (23) SacI (25) NheI (27) BmtI (31) AbsI - PaeR7I - PspXI - XhoI (33) MCS BglII (46) BglI - SfiI (59) HindIII (65) NcoI (98) BglI (100) BstAPI - DraIII (288) NsiI (319) PfoI * (380) XmnI (634) BsaI (640) NruI (761) ZraI (795) PflFI - Tth111I (796) AatII (797) PasI (874) EcoRI (1032) XbaI (1168) FseI (1187) AanI - PsiI (1308) HpaI (1328) MfeI (1337) BamHI (1430) SalI (1436) PshAI (1501) RVprimer4 (1487 .. 1506) BbsI (1515) AfeI (1562) BspQI - SapI (1570) AflIII - PciI (1686) pGL4.71 [hRlucP] 3653 bp
BglI  (8)
3 sites
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
SfiI  (8)
2 sites
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.
BsgI  (3651)
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  (3623)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Cleavage may be enhanced when more than one copy of the AarI recognition sequence is present.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its electrophoretic mobility.
BfuAI  (3623)
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  (3623)
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.
BsmBI  (3450)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different BsmBI sites may not be compatible.
BsmBI-v2 is an improved version of BsmBI.
SpeI  (3437)
1 site
A C T A G T T G A T C A
BstZ17I  (3118)
1 site
G T A T A C C A T A T G
SacII  (3002)
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.
PvuI  (2978)
1 site
C G A T C G G C T A G C
MscI  (2842)
1 site
T G G C C A A C C G G T
AhdI  (2608)
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.
BstEII  (2533)
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.
BstXI  (2530)
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.
PstI  (2530)
1 site
C T G C A G G A C G T C
AleI  (2528)
1 site
C A C N N N N G T G G T G N N N N C A C
NotI  (2506)
1 site
G C G G C C G C C G C C G G C G
BspHI  (2406)
1 site
T C A T G A A G T A C T
AlwNI  (2102)
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.
BaeGI  (2004)
1 site
G K G C M C C M C G K G

Sticky ends from different BaeGI sites may not be compatible.
Bme1580I  (2004)
1 site
G K G C M C C M C G K G

Sticky ends from different Bme1580I sites may not be compatible.
ApaLI  (2000)
1 site
G T G C A C C A C G T G
BciVI  (1889)
1 site
G T A T C C ( N ) 5 N C A T A G G ( N ) 5

The 1-base overhangs produced by BciVI may be hard to ligate.
Sticky ends from different BciVI sites may not be compatible.
BssSI  (1859)
1 site
C A C G A G G T G C T C
NspI  (1690)
1 site
R C A T G Y Y G T A C R
Acc65I  (14)
1 site
G G T A C C C C A T G G
KpnI  (18)
1 site
G G T A C C C C A T G G
Bpu10I  (19)
1 site
C C T N A G C G G A N T C G

Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
This recognition sequence is asymmetric, so ligating sticky ends generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
Eco53kI  (23)
1 site
G A G C T C C T C G A G
SacI  (25)
1 site
G A G C T C C T C G A G
NheI  (27)
1 site
G C T A G C C G A T C G
BmtI  (31)
1 site
G C T A G C C G A T C G
AbsI  (33)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (33)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (33)
1 site
V C T C G A G B B G A G C T C V
XhoI  (33)
1 site
C T C G A G G A G C T C
BglII  (46)
1 site
A G A T C T T C T A G A
BglI  (59)
3 sites
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
SfiI  (59)
2 sites
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.
HindIII  (65)
1 site
A A G C T T T T C G A A
NcoI  (98)
1 site
C C A T G G G G T A C C
BglI  (100)
3 sites
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
BstAPI  (288)
1 site
G C A N N N N N T G C C G T N N N N N A C G

Sticky ends from different BstAPI sites may not be compatible.
DraIII  (288)
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  (319)
1 site
A T G C A T T A C G T A
PfoI  (380)
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.
XmnI  (634)
1 site
G A A N N N N T T C C T T N N N N A A G
BsaI  (640)
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.
NruI  (761)
1 site
T C G C G A A G C G C T
ZraI  (795)
1 site
G A C G T C C T G C A G
PflFI  (796)
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  (796)
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.
AatII  (797)
1 site
G A C G T C C T G C A G
PasI  (874)
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.
EcoRI  (1032)
1 site
G A A T T C C T T A A G
XbaI  (1168)
1 site
T C T A G A A G A T C T
FseI  (1187)
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.
AanI  (1308)
1 site
T T A T A A A A T A T T
PsiI  (1308)
1 site
T T A T A A A A T A T T
HpaI  (1328)
1 site
G T T A A C C A A T T G
MfeI  (1337)
1 site
C A A T T G G T T A A C
BamHI  (1430)
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  (1436)
1 site
G T C G A C C A G C T G
PshAI  (1501)
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.
BbsI  (1515)
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.
AfeI  (1562)
1 site
A G C G C T T C G C G A
BspQI  (1570)
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  (1570)
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  (1686)
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  (1686)
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
3602 .. 3621  =  20 annealed bases
Tm  =  54°C
RVprimer4
20-mer  /  65% GC
1 binding site
1487 .. 1506  =  20 annealed bases
Tm  =  62°C
hRluc
100 .. 1032  =  933 bp
311 amino acids  =  36.0 kDa
Product: Renilla luciferase
human codon-optimized
hRluc
100 .. 1032  =  933 bp
311 amino acids  =  36.0 kDa
Product: Renilla luciferase
human codon-optimized
hPEST
1036 .. 1155  =  120 bp
40 amino acids  =  4.2 kDa
Product: PEST degradation sequence from mouse ornithine decarboxylase
human codon-optimized
hPEST
1036 .. 1155  =  120 bp
40 amino acids  =  4.2 kDa
Product: PEST degradation sequence from mouse ornithine decarboxylase
human codon-optimized
AmpR
2535 .. 3395  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   2535 .. 3326  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2535 .. 3395  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   3327 .. 3395  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2535 .. 3395  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
1747 .. 2335  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
1747 .. 2335  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
SV40 poly(A) signal
1207 .. 1328  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1207 .. 1328  =  122 bp
SV40 polyadenylation signal
pause site
3562 .. 3653  =  92 bp
RNA polymerase II transcriptional pause signal from the human α2 globin gene
pause site
3562 .. 3653  =  92 bp
RNA polymerase II transcriptional pause signal from the human α2 globin gene
MCS
1 .. 70  =  70 bp
multiple cloning site
MCS
1 .. 70  =  70 bp
multiple cloning site
poly(A) signal
3500 .. 3548  =  49 bp
synthetic polyadenylation signal
poly(A) signal
3500 .. 3548  =  49 bp
synthetic polyadenylation signal
ORF:  100 .. 1158  =  1059 bp
ORF:  352 amino acids  =  40.2 kDa
ORF:  2665 .. 2931  =  267 bp
ORF:  88 amino acids  =  9.3 kDa
ORF:  1293 .. 1535  =  243 bp
ORF:  80 amino acids  =  9.0 kDa
ORF:  2535 .. 3395  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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Download pGL4.71[hRlucP].dna file

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