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Plasmid Files

pSecTag/FRT/V5-His-TOPO® (linearized)

Linearized mammalian vector with 3'-T overhangs and bound topoisomerase, for TOPO® TA cloning of PCR products, FLP recombination, and secretion of tagged proteins.

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pSecTag FRT V5-His-TOPO (linearized).dna
Map and Sequence File:    Download    Open   
Sequence Author:  Thermo Fisher (Invitrogen)
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SgrDI (4171) SspI (4054) XmnI (3849) FspI (3472) AhdI (3250) PciI (2357) BspQI - SapI (2241) BstZ17I (1978) BsmI (1926) PsiI (1865) PfoI (1781) SacII (1471) BglII (4185) MfeI (4334) Bpu10I (4353) NruI (4381) SpeI (4422) SnaBI (4763) NheI (5068) BmtI (5072) SfiI (5150) MauBI (5160) BsiWI (5168) End (5185) Start (1) Acc65I (13) KpnI (17) BamHI (25) BseRI (46) AgeI (79) PmeI (109) BclI * (118) BbsI (320) SphI (331) XbaI (666) MscI (691) PshAI (718) BsmBI (759) EcoRI (934) BfuAI - BspMI (1004) PstI (1028) AsiSI (1055) RsrII (1099) pSecTag/FRT/V5-His-TOPO® 5184 bp
SgrDI  (4171)
1 site
C G T C G A C G G C A G C T G C
SspI  (4054)
1 site
A A T A T T T T A T A A
XmnI  (3849)
1 site
G A A N N N N T T C C T T N N N N A A G
FspI  (3472)
1 site
T G C G C A A C G C G T
AhdI  (3250)
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.
PciI  (2357)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (2241)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (2241)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be mixed before removing an aliquot.
BstZ17I  (1978)
1 site
G T A T A C C A T A T G
BsmI  (1926)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
PsiI  (1865)
1 site
T T A T A A A A T A T T
PfoI  (1781)
1 site
T C C N G G A A G G N C C T

Sticky ends from different PfoI sites may not be compatible.
SacII  (1471)
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.
BglII  (4185)
1 site
A G A T C T T C T A G A
MfeI  (4334)
1 site
C A A T T G G T T A A C
Bpu10I  (4353)
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.
NruI  (4381)
1 site
T C G C G A A G C G C T
SpeI  (4422)
1 site
A C T A G T T G A T C A
SnaBI  (4763)
1 site
T A C G T A A T G C A T
NheI  (5068)
1 site
G C T A G C C G A T C G
BmtI  (5072)
1 site
G C T A G C C G A T C G
SfiI  (5150)
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.
MauBI  (5160)
1 site
C G C G C G C G G C G C G C G C
BsiWI  (5168)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
End  (5185)
0 sites
Start  (1)
0 sites
Acc65I  (13)
1 site
G G T A C C C C A T G G
KpnI  (17)
1 site
G G T A C C C C A T G G
BamHI  (25)
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.
BseRI  (46)
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.
AgeI  (79)
1 site
A C C G G T T G G C C A
PmeI  (109)
1 site
G T T T A A A C C A A A T T T G
BclI  (118)
1 site
T G A T C A A C T A G T
* Blocked by Dam methylation.
BclI is typically used at 50-55°C, but is 50% active at 37°C.
BbsI  (320)
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.
SphI  (331)
1 site
G C A T G C C G T A C G
XbaI  (666)
1 site
T C T A G A A G A T C T
MscI  (691)
1 site
T G G C C A A C C G G T
PshAI  (718)
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.
BsmBI  (759)
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.
EcoRI  (934)
1 site
G A A T T C C T T A A G
BfuAI  (1004)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BfuAI recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (1004)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BspMI recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
PstI  (1028)
1 site
C T G C A G G A C G T C
AsiSI  (1055)
1 site
G C G A T C G C C G C T A G C G
RsrII  (1099)
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.
HygR
694 .. 1716  =  1023 bp
340 amino acids  =  37.9 kDa
Product: aminoglycoside phosphotransferase from E. coli
confers resistance to hygromycin
HygR
694 .. 1716  =  1023 bp
340 amino acids  =  37.9 kDa
Product: aminoglycoside phosphotransferase from E. coli
confers resistance to hygromycin
AmpR
3177 .. 4037  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   3177 .. 3968  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3177 .. 4037  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   3969 .. 4037  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3177 .. 4037  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
2418 .. 3006  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication