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pSF-CMV-Ub-FLuc AscI

Mammalian vector for co-expressing a gene together with firefly luciferase expressed from the ubiquitin C promoter.

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pSF-CMV-Ub-FLuc AscI Sequence and MappSF-CMV-Ub-FLuc AscI.dna
Map and Sequence File   
Sequence Author:  Oxford Genetics
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 PmeI (7081) RsrII (6774) BsrDI (6491) PflFI - Tth111I (6376) PmeI (6033) AscI (5876) SexAI * (5481) AgeI (5273) BlpI (5096) FspI (4964) AleI (4640) BsrGI (4632) ScaI (4307) PvuII (4216) EcoNI (3869) AvrII (3865) AarI (3819) AanI (3792) AsiSI - PvuI (5) BglII (232) SnaBI (566) CMV promoter 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) BamHI (986) StuI (996) NheI (1002) BmtI (1006) stop codons AanI (1152) MfeI (1181) T7 terminator PstI - SbfI (1518) 3' β- globin insulator BstEII (1624) PacI (1650) SwaI (1776) AlwNI (2196) SwaI (2669) FseI (2775) AscI (2921) SacII (2959) BsaI (3394) pSF-CMV-Ub-FLuc AscI 7200 bp
PmeI  (7081)
2 sites
G T T T A A A C C A A A T T T G
RsrII  (6774)
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.
BsrDI  (6491)
1 site
G C A A T G N N C G T T A C

Sticky ends from different BsrDI sites may not be compatible.
PflFI  (6376)
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  (6376)
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  (6033)
2 sites
G T T T A A A C C A A A T T T G
AscI  (5876)
2 sites
G G C G C G C C C C G C G C G G
SexAI  (5481)
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.
AgeI  (5273)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
BlpI  (5096)
1 site
G C T N A G C C G A N T C G

Sticky ends from different BlpI sites may not be compatible.
FspI  (4964)
1 site
T G C G C A A C G C G T
AleI  (4640)
1 site
C A C N N N N G T G G T G N N N N C A C
BsrGI  (4632)
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.
ScaI  (4307)
1 site
A G T A C T T C A T G A
PvuII  (4216)
1 site
C A G C T G G T C G A C
EcoNI  (3869)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
AvrII  (3865)
1 site
C C T A G G G G A T C C
AarI  (3819)
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.
AanI  (3792)
2 sites
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
BamHI  (986)
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.
StuI  (996)
1 site
A G G C C T T C C G G A
NheI  (1002)
1 site
G C T A G C C G A T C G
BmtI  (1006)
1 site
G C T A G C C G A T C G
AanI  (1152)
2 sites
T T A T A A A A T A T T
MfeI  (1181)
1 site
C A A T T G G T T A A C
PstI  (1518)
1 site
C T G C A G G A C G T C
SbfI  (1518)
1 site
C C T G C A G G G G A C G T C C
BstEII  (1624)
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.
PacI  (1650)
1 site
T T A A T T A A A A T T A A T T
SwaI  (1776)
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.
AlwNI  (2196)
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.
SwaI  (2669)
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.
FseI  (2775)
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.
AscI  (2921)
2 sites
G G C G C G C C C C G C G C G G
SacII  (2959)
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.
BsaI  (3394)
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.
luciferase
4142 .. 5794  =  1653 bp
550 amino acids  =  60.6 kDa
Product: firefly luciferase
synthetic luc2 version of the luciferase gene
luciferase
4142 .. 5794  =  1653 bp
550 amino acids  =  60.6 kDa
Product: firefly luciferase
synthetic luc2 version of the luciferase gene
UbC promoter
2929 .. 4138  =  1210 bp
human ubiquitin C promoter
UbC promoter
2929 .. 4138  =  1210 bp
human ubiquitin C promoter
NeoR/KanR
6130 .. 6924  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
6130 .. 6924  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
ori
1841 .. 2429  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1841 .. 2429  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
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
1363 .. 1499  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
1363 .. 1499  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
6937 .. 7073  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
6937 .. 7073  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
SV40 poly(A) signal
1051 .. 1172  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1051 .. 1172  =  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
1544 .. 1615  =  72 bp
insulator downstream of the human β-globin locus
(Farrell et al., 2002)
3' β-globin insulator
1544 .. 1615  =  72 bp
insulator downstream of the human β-globin locus
(Farrell et al., 2002)
poly(A) signal
5816 .. 5864  =  49 bp
synthetic polyadenylation signal
poly(A) signal
5816 .. 5864  =  49 bp
synthetic polyadenylation signal
T7 terminator
1287 .. 1333  =  47 bp
transcription terminator for bacteriophage T7 RNA
polymerase
T7 terminator
1287 .. 1333  =  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
1009 .. 1019  =  11 bp
stop codons in all three reading frames
stop codons
1009 .. 1019  =  11 bp
stop codons in all three reading frames
RBS
6117 .. 6122  =  6 bp
Shine-Dalgarno ribosome binding site
RBS
6117 .. 6122  =  6 bp
Shine-Dalgarno ribosome binding site
Kozak sequence
903 .. 909  =  7 bp
Kozak sequence
903 .. 909  =  7 bp
RBS
893 .. 898  =  6 bp
Shine-Dalgarno ribosome binding site
RBS
893 .. 898  =  6 bp
Shine-Dalgarno ribosome binding site
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