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Plasmid Files

pF5K CMV-neo

Flexi® vector with a kanamycin/G418 resistance marker, for protein expression in mammalian cells.

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pF5K CMV-neo Sequence and MappF5K CMV-neo.dna
Map and Sequence File   
Sequence Author:  Promega
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 ScaI (4554) MluI (4470) BspEI (4257) ApaLI (3807) PciI (3493) BspDI - ClaI (3318) BstXI (3258) BstBI (3219) RsrII (3053) NaeI (3039) NgoMIV (3037) BssHII (2934) PflFI - Tth111I (2655) PluTI (2540) SfoI (2538) NarI (2537) KasI (2536) PacI (2324) AvrII (2300) StuI (2299) BglII (4589) BsrGI (96) SpeI (152) NdeI (387) SnaBI (493) BsaI (882) BbsI (928) T7 promoter NheI (1052) BmtI (1056) AsiSI - PvuI - SgfI (1060) DraI - PmeI (1427) EcoRI (1432) XbaI (1459) SalI (1465) AccI (1466) SbfI (1475) BlpI (1540) NotI (1570) Bts α I (1639) PsiI (1693) HpaI (1713) MfeI (1722) SexAI * (2067) SfiI (2253) BseRI (2296) pF5K CMV-neo 4594 bp
ScaI  (4554)
1 site
A G T A C T T C A T G A
MluI  (4470)
1 site
A C G C G T T G C G C A
BspEI  (4257)
1 site
T C C G G A A G G C C T
ApaLI  (3807)
1 site
G T G C A C C A C G T G
PciI  (3493)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspDI  (3318)
1 site
A T C G A T T A G C T A
ClaI  (3318)
1 site
A T C G A T T A G C T A
BstXI  (3258)
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.
BstBI  (3219)
1 site
T T C G A A A A G C T T
RsrII  (3053)
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.
NaeI  (3039)
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  (3037)
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.
BssHII  (2934)
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.
PflFI  (2655)
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  (2655)
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.
PluTI  (2540)
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.
SfoI  (2538)
1 site
G G C G C C C C G C G G
NarI  (2537)
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.
KasI  (2536)
1 site
G G C G C C C C G C G G
PacI  (2324)
1 site
T T A A T T A A A A T T A A T T
AvrII  (2300)
1 site
C C T A G G G G A T C C
StuI  (2299)
1 site
A G G C C T T C C G G A
BglII  (4589)
1 site
A G A T C T T C T A G A
BsrGI  (96)
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.
SpeI  (152)
1 site
A C T A G T T G A T C A
NdeI  (387)
1 site
C A T A T G G T A T A C

Prolonged incubation with NdeI may lead to removal of additional
nucleotides.
SnaBI  (493)
1 site
T A C G T A A T G C A T
BsaI  (882)
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.
BbsI  (928)
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.
NheI  (1052)
1 site
G C T A G C C G A T C G
BmtI  (1056)
1 site
G C T A G C C G A T C G
AsiSI  (1060)
1 site
G C G A T C G C C G C T A G C G
PvuI  (1060)
1 site
C G A T C G G C T A G C
SgfI  (1060)
1 site
G C G A T C G C C G C T A G C G
DraI  (1427)
1 site
T T T A A A A A A T T T
PmeI  (1427)
1 site
G T T T A A A C C A A A T T T G
EcoRI  (1432)
1 site
G A A T T C C T T A A G
XbaI  (1459)
1 site
T C T A G A A G A T C T
SalI  (1465)
1 site
G T C G A C C A G C T G
AccI  (1466)
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.
SbfI  (1475)
1 site
C C T G C A G G G G A C G T C C
BlpI  (1540)
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.
NotI  (1570)
1 site
G C G G C C G C C G C C G G C G
BtsαI  (1639)
1 site
G C A G T G N N C G T C A C

Sticky ends from different BtsI sites may not be compatible.
PsiI  (1693)
1 site
T T A T A A A A T A T T
HpaI  (1713)
1 site
G T T A A C C A A T T G
MfeI  (1722)
1 site
C A A T T G G T T A A C
SexAI  (2067)
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.
SfiI  (2253)
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.
BseRI  (2296)
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.
NeoR/KanR
2409 .. 3203  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
2409 .. 3203  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
ori
3554 .. 4142  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3554 .. 4142  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
CMV enhancer
138 .. 517  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
138 .. 517  =  380 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
1958 .. 2315  =  358 bp
SV40 enhancer and early promoter
SV40 promoter
1958 .. 2315  =  358 bp
SV40 enhancer and early promoter
barnase
1087 .. 1422  =  336 bp
111 amino acids  =  12.5 kDa
Product: ribonuclease from Bacillus
amyloliquefaciens

The barnase gene is lethal in standard bacterial
transformation strains.
barnase
1087 .. 1422  =  336 bp
111 amino acids  =  12.5 kDa
Product: ribonuclease from Bacillus
amyloliquefaciens

The barnase gene is lethal in standard bacterial
transformation strains.
cer region
4258 .. 4541  =  284 bp
ColE1-derived recombination site that helps to
maintain plasmids as monomers
cer region
4258 .. 4541  =  284 bp
ColE1-derived recombination site that helps to
maintain plasmids as monomers
CMV promoter
518 .. 721  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
518 .. 721  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
chimeric intron
857 .. 989  =  133 bp
chimera between introns from human β-globin and
immunoglobulin heavy chain genes
chimeric intron
857 .. 989  =  133 bp
chimera between introns from human β-globin and
immunoglobulin heavy chain genes
SV40 poly(A) signal
1592 .. 1713  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1592 .. 1713  =  122 bp
SV40 polyadenylation signal
poly(A) signal
3267 .. 3315  =  49 bp
synthetic polyadenylation signal
poly(A) signal
3267 .. 3315  =  49 bp
synthetic polyadenylation signal
EM7 promoter
2330 .. 2377  =  48 bp
synthetic bacterial promoter
EM7 promoter
2330 .. 2377  =  48 bp
synthetic bacterial promoter
T7 promoter
1033 .. 1051  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
1033 .. 1051  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SV40 ori
2166 .. 2301  =  136 bp
SV40 origin of replication
SV40 ori
2166 .. 2301  =  136 bp
SV40 origin of replication
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