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Plasmid Files

pSF-CMVe-RLuc

Vector encoding the CMV enhancer and Renilla luciferase, for measuring the activity of promoters.

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pSF-CMVe-RLuc Sequence and MappSF-CMVe-RLuc.dna
Map and Sequence File   
Sequence Author:  Oxford Genetics
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 PmeI (5079) RsrII (4772) BsrDI (4489) PflFI - Tth111I (4374) PmeI (4031) PpuMI - SanDI (3965) AscI (3874) FseI (3728) NaeI (3726) NgoMIV (3724) SwaI (3622) BspHI (3453) AlwNI (3149) SwaI (2729) PacI (2603) AsiSI - PvuI (5) BglII (232) AseI (244) CMV enhancer SnaBI (577) NruI (710) PshAI (716) ScaI (728) AarI (793) SalI (824) AccI (825) SpeI (832) TspMI - XmaI (840) SmaI (842) PspOMI (848) ApaI (852) BstBI (857) BglII (865) EagI - NotI (907) HindIII (918) Eco53kI (928) SacI (930) EcoRI (934) Acc65I (946) KpnI (950) BstAPI (1144) AanI (1248) Bpu10I (1604) BseRI - BsgI (1893) stop codons BspDI - ClaI (1930) BamHI (1939) StuI (1949) NheI (1955) BmtI (1959) stop codons AanI (2105) HpaI (2125) MfeI (2134) PstI - SbfI (2471) 3' β- globin insulator BstEII (2577) pSF-CMVe-RLuc 5198 bp
PmeI  (5079)
2 sites
G T T T A A A C C A A A T T T G
RsrII  (4772)
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  (4489)
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  (4374)
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  (4374)
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  (4031)
2 sites
G T T T A A A C C A A A T T T G
PpuMI  (3965)
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.
SanDI  (3965)
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  (3874)
1 site
G G C G C G C C C C G C G C G G
FseI  (3728)
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.
NaeI  (3726)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI
recognition sequence.
NgoMIV  (3724)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV
recognition sequence.
SwaI  (3622)
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.
BspHI  (3453)
1 site
T C A T G A A G T A C T
AlwNI  (3149)
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  (2729)
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  (2603)
1 site
T T A A T T A A A A T T A 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
AseI  (244)
1 site
A T T A A T T A A T T A
SnaBI  (577)
1 site
T A C G T A A T G C A T
NruI  (710)
1 site
T C G C G A A G C G C T
PshAI  (716)
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.
ScaI  (728)
1 site
A G T A C T T C A T G A
AarI  (793)
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.
SalI  (824)
1 site
G T C G A C C A G C T G
AccI  (825)
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.
SpeI  (832)
1 site
A C T A G T T G A T C A
TspMI  (840)
1 site
C C C G G G G G G C C C
XmaI  (840)
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  (842)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
PspOMI  (848)
1 site
G G G C C C C C C G G G
ApaI  (852)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BstBI  (857)
1 site
T T C G A A A A G C T T
BglII  (865)
2 sites
A G A T C T T C T A G A
EagI  (907)
1 site
C G G C C G G C C G G C
NotI  (907)
1 site
G C G G C C G C C G C C G G C G
HindIII  (918)
1 site
A A G C T T T T C G A A
Eco53kI  (928)
1 site
G A G C T C C T C G A G
SacI  (930)
1 site
G A G C T C C T C G A G
EcoRI  (934)
1 site
G A A T T C C T T A A G
Acc65I  (946)
1 site
G G T A C C C C A T G G
KpnI  (950)
1 site
G G T A C C C C A T G G
BstAPI  (1144)
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.
AanI  (1248)
2 sites
T T A T A A A A T A T T
Bpu10I  (1604)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
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.
BseRI  (1893)
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  (1893)
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  (1930)
1 site
A T C G A T T A G C T A
ClaI  (1930)
1 site
A T C G A T T A G C T A
BamHI  (1939)
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  (1949)
1 site
A G G C C T T C C G G A
NheI  (1955)
1 site
G C T A G C C G A T C G
BmtI  (1959)
1 site
G C T A G C C G A T C G
AanI  (2105)
2 sites
T T A T A A A A T A T T
HpaI  (2125)
1 site
G T T A A C C A A T T G
MfeI  (2134)
1 site
C A A T T G G T T A A C
PstI  (2471)
1 site
C T G C A G G A C G T C
SbfI  (2471)
1 site
C C T G C A G G G G A C G T C C
BstEII  (2577)
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.
RLuc
956 .. 1894  =  939 bp
312 amino acids  =  36.1 kDa
Product: Renilla luciferase
codon-optimized synthetic gene
RLuc
956 .. 1894  =  939 bp
312 amino acids  =  36.1 kDa
Product: Renilla luciferase
codon-optimized synthetic gene
NeoR/KanR
4128 .. 4922  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
4128 .. 4922  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
ori
2794 .. 3382  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2794 .. 3382  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
CMV enhancer
298 .. 601  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
298 .. 601  =  304 bp
human cytomegalovirus immediate early enhancer
rrnG terminator
2316 .. 2452  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
2316 .. 2452  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
4935 .. 5071  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
rrnG terminator
4935 .. 5071  =  137 bp
transcription terminator from the E. coli ribosomal
RNA rrnG operon (Albrechtsen et al., 1991)
SV40 poly(A) signal
2004 .. 2125  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2004 .. 2125  =  122 bp
SV40 polyadenylation signal
pause site
732 .. 823  =  92 bp
RNA polymerase II transcriptional pause signal from
the human α2 globin gene
pause site
732 .. 823  =  92 bp
RNA polymerase II transcriptional pause signal from
the human α2 globin gene
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
2497 .. 2568  =  72 bp
insulator downstream of the human β-globin locus
(Farrell et al., 2002)
3' β-globin insulator
2497 .. 2568  =  72 bp
insulator downstream of the human β-globin locus
(Farrell et al., 2002)
T7 terminator
2240 .. 2286  =  47 bp
transcription terminator for bacteriophage T7 RNA
polymerase
T7 terminator
2240 .. 2286  =  47 bp
transcription terminator for bacteriophage T7 RNA
polymerase
MCS
824 .. 861  =  38 bp
multiple cloning site
MCS
824 .. 861  =  38 bp
multiple cloning site
stop codons
1914 .. 1924  =  11 bp
stop codons in all three reading frames
stop codons
1914 .. 1924  =  11 bp
stop codons in all three reading frames
stop codons
1962 .. 1972  =  11 bp
stop codons in all three reading frames
stop codons
1962 .. 1972  =  11 bp
stop codons in all three reading frames
RBS
942 .. 947  =  6 bp
Shine-Dalgarno ribosome binding site
RBS
942 .. 947  =  6 bp
Shine-Dalgarno ribosome binding site
RBS
4115 .. 4120  =  6 bp
Shine-Dalgarno ribosome binding site
RBS
4115 .. 4120  =  6 bp
Shine-Dalgarno ribosome binding site
Kozak sequence
952 .. 958  =  7 bp
Kozak sequence
952 .. 958  =  7 bp
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