pF5A CMV-neo

Flexi® vector with a ampicillin and G418 resistance markers, for protein expression in mammalian cells.

Sequence Author: Promega

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BglII (5586) MluI (5467) BspEI (5254) PciI (4490) AgeI (4422) AhdI (4320) BpmI (4251) XmnI (3720) BspDI - ClaI (3275) BstXI (3215) RsrII (3010) NaeI (2996) NgoMIV (2994) BssHII (2891) BsrGI (96) SpeI (152) NdeI (387) 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) PsiI (1693) HpaI (1713) MfeI (1722) SexAI * (2067) SfiI (2253) BseRI (2296) StuI (2299) AvrII (2300) KasI (2493) NarI (2494) SfoI (2495) PluTI (2497) PflFI - Tth111I (2612) pF5A CMV-neo 5591 bp
BglII  (5586)
1 site
A G A T C T T C T A G A
MluI  (5467)
1 site
A C G C G T T G C G C A
BspEI  (5254)
1 site
T C C G G A A G G C C T
PciI  (4490)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AgeI  (4422)
1 site
A C C G G T T G G C C A
AhdI  (4320)
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.
BpmI  (4251)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its electrophoretic mobility.
BpmI quickly loses activity at 37°C.
XmnI  (3720)
1 site
G A A N N N N T T C C T T N N N N A A G
BspDI  (3275)
1 site
A T C G A T T A G C T A
ClaI  (3275)
1 site
A T C G A T T A G C T A
BstXI  (3215)
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.
RsrII  (3010)
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  (2996)
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  (2994)
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  (2891)
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.
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.
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
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.
StuI  (2299)
1 site
A G G C C T T C C G G A
AvrII  (2300)
1 site
C C T A G G G G A T C C
KasI  (2493)
1 site
G G C G C C C C G C G G
NarI  (2494)
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.
SfoI  (2495)
1 site
G G C G C C C C G C G G
PluTI  (2497)
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.
PflFI  (2612)
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  (2612)
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.
AmpR
3533 .. 4393  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   3533 .. 3601  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3533 .. 4393  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   3602 .. 4393  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3533 .. 4393  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
NeoR/KanR
2366 .. 3160  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
2366 .. 3160  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
ori
4551 .. 5139  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
4551 .. 5139  =  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
5255 .. 5538  =  284 bp
ColE1-derived recombination site that helps to maintain plasmids as monomers
cer region
5255 .. 5538  =  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
AmpR promoter
3428 .. 3532  =  105 bp
AmpR promoter
3428 .. 3532  =  105 bp
poly(A) signal
3224 .. 3272  =  49 bp
synthetic polyadenylation signal
poly(A) signal
3224 .. 3272  =  49 bp
synthetic polyadenylation signal
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
ORF:  1087 .. 1422  =  336 bp
ORF:  111 amino acids  =  12.5 kDa
ORF:  2366 .. 3160  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  3533 .. 4393  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  2538 .. 2924  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  3036 .. 3341  =  306 bp
ORF:  101 amino acids  =  11.5 kDa
ORF:  821 .. 1066  =  246 bp
ORF:  81 amino acids  =  8.6 kDa
ORF:  2675 .. 3211  =  537 bp
ORF:  178 amino acids  =  19.8 kDa
ORF:  3997 .. 4263  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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