pEXP5-NT TOPO (linearized)

Linearized vector with 3'-T overhangs and bound topoisomerase, for TOPO® TA cloning and inducible bacterial expression of an N-terminally 6xHis-tagged protein.

Sequence Author: Thermo Fisher (Invitrogen)

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HomePlasmidsTA and GC Cloning VectorspEXP5-NT TOPO (linearized)
BglII (2581) AvaI - BsoBI (2577) MscI * (2560) BtgI (2555) FspAI (2550) Bpu10I (2422) BsgI * (2373) BtgZI (2343) BstZ17I (2258) AccI (2257) BspQI - SapI (2145) NspI (2032) AflIII - PciI (2028) DrdI (1926) PspFI (1728) BseYI (1724) AlwNI (1619) BanI (1187) XbaI (2639) ATG End (2745) Start (1) BlpI (59) StyI (81) ZraI (219) AatII (221) SspI (335) XmnI (540) HincII (598) ScaI (659) PvuI (771) NmeAIII (993) BglI (1022) BsrFI (1055) BpmI (1071) BmrI (1100) AhdI (1140) pEXP5-NT/TOPO® 2744 bp
BglII  (2581)
1 site		 A G A T C T T C T A G A
AvaI  (2577)
1 site		 C Y C G R G G R G C Y C
Sticky ends from different AvaI sites may not be compatible.
BsoBI  (2577)
1 site		 C Y C G R G G R G C Y C
Sticky ends from different BsoBI sites may not be compatible.
BsoBI is typically used at 37°C, but can be used at temperatures up to 65°C.
MscI  (2560)
1 site		 T G G C C A A C C G G T
* Blocked by Dcm methylation.
BtgI  (2555)
1 site		 C C R Y G G G G Y R C C
Sticky ends from different BtgI sites may not be compatible.
FspAI  (2550)
1 site		 R T G C G C A Y Y A C G C G T R
Bpu10I  (2422)
1 site		 C C T N A G C G G A N T C G
Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
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.
BsgI  (2373)
1 site		 G T G C A G ( N ) 14 N N C A C G T C ( N ) 14
* Blocked by EcoKI methylation.
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).
BtgZI  (2343)
1 site		 G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4
Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
BstZ17I  (2258)
1 site		 G T A T A C C A T A T G
AccI  (2257)
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.
BspQI  (2145)
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  (2145)
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.
NspI  (2032)
1 site		 R C A T G Y Y G T A C R
AflIII  (2028)
1 site		 A C R Y G T T G Y R C A
Sticky ends from different AflIII sites may not be compatible.
PciI  (2028)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
DrdI  (1926)
1 site		 G A C N N N N N N G T C C T G N N N N N N C A G
Sticky ends from different DrdI sites may not be compatible.
PspFI  (1728)
1 site		 C C C A G C G G G T C G
BseYI  (1724)
1 site		 C C C A G C G G G T C G
After cleavage, BseYI can remain bound to DNA and alter its electrophoretic mobility.
AlwNI  (1619)
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.
BanI  (1187)
1 site		 G G Y R C C C C R Y G G
Sticky ends from different BanI sites may not be compatible.
XbaI  (2639)
1 site		 T C T A G A A G A T C T
End  (2745)
0 sites
Start  (1)
0 sites
BlpI  (59)
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.
StyI  (81)
1 site		 C C W W G G G G W W C C
Sticky ends from different StyI sites may not be compatible.
ZraI  (219)
1 site		 G A C G T C C T G C A G
AatII  (221)
1 site		 G A C G T C C T G C A G
SspI  (335)
1 site		 A A T A T T T T A T A A
XmnI  (540)
1 site		 G A A N N N N T T C C T T N N N N A A G
HincII  (598)
1 site		 G T Y R A C C A R Y T G
ScaI  (659)
1 site		 A G T A C T T C A T G A
PvuI  (771)
1 site		 C G A T C G G C T A G C
NmeAIII  (993)
1 site		 G C C G A G ( N ) 18-19 N N C G G C T C ( N ) 18-19
Efficient cleavage requires at least two copies of the NmeAIII recognition sequence.
Sticky ends from different NmeAIII sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
BglI  (1022)
1 site		 G C C N N N N N G G C C G G N N N N N C C G
Sticky ends from different BglI sites may not be compatible.
BsrFI  (1055)
1 site		 R C C G G Y Y G G C C R
Cleavage may be enhanced when more than one copy of the BsrFI recognition sequence is present.
After cleavage, BsrFI can remain bound to DNA and alter its electrophoretic mobility.
BpmI  (1071)
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.
BmrI  (1100)
1 site		 A C T G G G ( N ) 4 N T G A C C C ( N ) 4
The 1-base overhangs produced by BmrI may be hard to ligate.
Sticky ends from different BmrI sites may not be compatible.
Unlike most restriction enzymes, BmrI can cleave DNA in the absence of magnesium.
AhdI  (1140)
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.
AmpR
353 .. 1213  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
   Segment 1:  signal sequence  
   353 .. 421  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
353 .. 1213  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
   Segment 2:  
   422 .. 1213  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
353 .. 1213  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
1384 .. 1972  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
ori
1384 .. 1972  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
AmpR promoter
248 .. 352  =  105 bp
AmpR promoter
248 .. 352  =  105 bp
ATG
2680 .. 2682  =  3 bp
1 amino acid  =  149.2 Da
ATG
2680 .. 2682  =  3 bp
1 amino acid  =  149.2 Da
6xHis
2692 .. 2709  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
2692 .. 2709  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
TEV site
2722 .. 2742  =  21 bp
7 amino acids  =  900.0 Da
Product: tobacco etch virus (TEV) recognition and cleavage site
TEV site
2722 .. 2742  =  21 bp
7 amino acids  =  900.0 Da
Product: tobacco etch virus (TEV) recognition and cleavage site
T7 terminator
70 .. 117  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
70 .. 117  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 promoter
2601 .. 2619  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
2601 .. 2619  =  19 bp
promoter for bacteriophage T7 RNA polymerase
RBS
2668 .. 2673  =  6 bp
ribosome binding site
RBS
2668 .. 2673  =  6 bp
ribosome binding site
HisG epitope
2692 .. 2712  =  21 bp
Product: HHHHHHG epitope
HisG epitope
2692 .. 2712  =  21 bp
Product: HHHHHHG epitope
ORF:  353 .. 1213  =  861 bp
ORF:  286 amino acids  =  31.5 kDa
ORF:  817 .. 1083  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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Individual Sequences & Maps

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pTZ57R_T
pcDNA5 FRT TO-TOPO (linearized)
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RBC T&A Cloning Vector (linearized)
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pSMART GC HK (linearized)
T-Vector pMD19 (Simple)
pcDNA6.2 N-EmGFP-GW TOPO (linearized)
pGlow-TOPO (linearized)
pSMART GC LK (linearized)
T-Vector pMD20
pCR-XL-TOPO (linearized)
pIB V5-His-TOPO (linearized)

Plasmid Sets

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