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Plasmid Files

pSF-CMV-PGK-FLuc

Dual-promoter mammalian vector for strong constitutive expression of a gene together with firefly luciferase.

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pSF-CMV-PGK-FLuc Sequence and MappSF-CMV-PGK-FLuc.dna
Map and Sequence File   
Sequence Author:  Oxford Genetics
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 PmeI (6275) RsrII (5968) PflFI - Tth111I (5570) PmeI (5227) SanDI (5161) AscI - BssHII (5070) FseI (4924) SwaI (4818) ApaLI (4243) SwaI (3925) PacI (3799) BstEII (3773) 3' β- globin insulator PstI - SbfI (3667) MfeI (3330) AanI - AanI - PsiI (3301) stop codons AsiSI - PvuI (5) BglII (232) CMV enhancer SnaBI (566) BglII (816) EagI - NotI (858) HindIII (869) Eco53kI (879) SacI (881) EcoRI (885) KpnI (901) NcoI (905) KpnI (915) EcoRV (921) AbsI - PaeR7I - PspXI - XhoI (928) XbaI (937) BseRI - BsgI (940) stop codons BspDI - ClaI (977) SphI (1027) BsmBI - SpeI (1233) BspEI (1407) BamHI (1488) BbsI (1556) PvuII (1572) ScaI (1663) BsrGI (1988) AleI (1996) FspI (2320) DraIII (2644) SexAI * (2837) NheI (3151) BmtI (3155) pSF-CMV-PGK-FLuc 6394 bp
PmeI  (6275)
2 sites
G T T T A A A C C A A A T T T G
RsrII  (5968)
1 site
C G G W C C G G C C W G G C

Efficient cleavage requires at least two copies of the RsrII
recognition sequence.
Sticky ends from different RsrII sites may not be compatible.
For full activity, add fresh DTT.
PflFI  (5570)
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  (5570)
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.
PmeI  (5227)
2 sites
G T T T A A A C C A A A T T T G
SanDI  (5161)
1 site
G G G W C C C C C C W G G G

Sticky ends from different SanDI sites may not be compatible.
AscI  (5070)
1 site
G G C G C G C C C C G C G C G G
BssHII  (5070)
1 site
G C G C G C C G C G C G

BssHII is typically used at 50°C, but is 75% active at 37°C.
FseI  (4924)
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.
SwaI  (4818)
2 sites
A T T T A A A T T A A A T T T A

SwaI is typically used at 25°C, but is 50% active at 37°C.
ApaLI  (4243)
1 site
G T G C A C C A C G T G
SwaI  (3925)
2 sites
A T T T A A A T T A A A T T T A

SwaI is typically used at 25°C, but is 50% active at 37°C.
PacI  (3799)
1 site
T T A A T T A A A A T T A A T T
BstEII  (3773)
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.
PstI  (3667)
1 site
C T G C A G G A C G T C
SbfI  (3667)
1 site
C C T G C A G G G G A C G T C C
MfeI  (3330)
1 site
C A A T T G G T T A A C
AanI  (3301)
1 site
T T A T A A A A T A T T
AanI  (3301)
1 site
T T A T A A A A T A T T
PsiI  (3301)
1 site
T T A T A A A A T A T T
AsiSI  (5)
1 site
G C G A T C G C C G C T A G C G
PvuI  (5)
1 site
C G A T C G G C T A G C
BglII  (232)
2 sites
A G A T C T T C T A G A
SnaBI  (566)
1 site
T A C G T A A T G C A T
BglII  (816)
2 sites
A G A T C T T C T A G A
EagI  (858)
1 site
C G G C C G G C C G G C
NotI  (858)
1 site
G C G G C C G C C G C C G G C G
HindIII  (869)
1 site
A A G C T T T T C G A A
Eco53kI  (879)
1 site
G A G C T C C T C G A G
SacI  (881)
1 site
G A G C T C C T C G A G
EcoRI  (885)
1 site
G A A T T C C T T A A G
KpnI  (901)
2 sites
G G T A C C C C A T G G
NcoI  (905)
1 site
C C A T G G G G T A C C
KpnI  (915)
2 sites
G G T A C C C C A T G G
EcoRV  (921)
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.
AbsI  (928)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (928)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (928)
1 site
V C T C G A G B B G A G C T C V
XhoI  (928)
1 site
C T C G A G G A G C T C
XbaI  (937)
1 site
T C T A G A A G A T C T
BseRI  (940)
1 site
G A G G A G ( N ) 8 N N C T C C T C ( N ) 8

Sticky ends from different BseRI sites may not be compatible.
BseRI quickly loses activity at 37°C.
Prolonged incubation with BseRI may lead to degradation of the
DNA.
BsgI  (940)
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).
BspDI  (977)
1 site
A T C G A T T A G C T A
ClaI  (977)
1 site
A T C G A T T A G C T A
SphI  (1027)
1 site
G C A T G C C G T A C G
BsmBI  (1233)
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.
SpeI  (1233)
1 site
A C T A G T T G A T C A
BspEI  (1407)
1 site
T C C G G A A G G C C T
BamHI  (1488)
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.
BbsI  (1556)
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.
PvuII  (1572)
1 site
C A G C T G G T C G A C
ScaI  (1663)
1 site
A G T A C T T C A T G A
BsrGI  (1988)
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.
AleI  (1996)
1 site
C A C N N N N G T G G T G N N N N C A C
FspI  (2320)
1 site
T G C G C A A C G C G T
DraIII  (2644)
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.
SexAI  (2837)
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.
NheI  (3151)
1 site
G C T A G C C G A T C G
BmtI  (3155)
1 site
G C T A G C C G A T C G
luciferase
1498 .. 3150  =  1653 bp
550 amino acids  =  60.6 kDa
Product: firefly luciferase
synthetic luc2 version of the luciferase gene
luciferase
1498 .. 3150  =  1653 bp
550 amino acids  =  60.6 kDa
Product: firefly luciferase
synthetic luc2 version of the luciferase gene
NeoR/KanR
5324 .. 6118  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
5324 .. 6118  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
ori
3990 .. 4578  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3990 .. 4578  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
PGK promoter
988 .. 1487  =  500 bp
mouse phosphoglycerate kinase 1 promoter
PGK promoter
988 .. 1487  =  500 bp
mouse phosphoglycerate kinase 1 promoter
CMV enhancer
287 .. 590  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
287 .. 590  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
591 .. 794  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
591 .. 794  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
rrnG terminator
3512 .. 3648  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
3512 .. 3648  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
6131 .. 6267  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
6131 .. 6267  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
SV40 poly(A) signal
3200 .. 3321  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
3200 .. 3321  =  122 bp
SV40 polyadenylation signal
MCS
857 .. 942  =  86 bp
multiple cloning site
MCS
857 .. 942  =  86 bp
multiple cloning site
5' β-globin insulator
18 .. 89  =  72 bp
insulator upstream of the human β-globin locus
(Farrell et al., 2002)
5' β-globin insulator
18 .. 89  =  72 bp
insulator upstream of the human β-globin locus
(Farrell et al., 2002)
3' β-globin insulator
3693 .. 3764  =  72 bp
insulator downstream of the human β-globin locus
(Farrell et al., 2002)
3' β-globin insulator
3693 .. 3764  =  72 bp
insulator downstream of the human β-globin locus
(Farrell et al., 2002)
T7 terminator
3436 .. 3482  =  47 bp
transcription terminator for bacteriophage T7 RNA
polymerase
T7 terminator
3436 .. 3482  =  47 bp
transcription terminator for bacteriophage T7 RNA
polymerase
stop codons
961 .. 971  =  11 bp
stop codons in all three reading frames
stop codons
961 .. 971  =  11 bp
stop codons in all three reading frames
stop codons
3158 .. 3168  =  11 bp
stop codons in all three reading frames
stop codons
3158 .. 3168  =  11 bp
stop codons in all three reading frames
RBS
5311 .. 5316  =  6 bp
Shine-Dalgarno ribosome binding site
RBS
5311 .. 5316  =  6 bp
Shine-Dalgarno ribosome binding site
Kozak sequence
903 .. 909  =  7 bp
Kozak sequence
903 .. 909  =  7 bp
RBS
893 .. 898  =  6 bp
Shinbe-Dalgarno ribosome binding site
RBS
893 .. 898  =  6 bp
Shinbe-Dalgarno ribosome binding site
ATG
907 .. 909  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
907 .. 909  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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