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Plasmid Files

pEXP1-DEST

Gateway® destination vector for cell-free expression and N-terminal tagging.

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pEXP1-DEST Sequence and MappEXP1-DEST.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 BspQI - SapI (4560) NspI (4447) AflIII - PciI (4443) PspFI (4143) BseYI (4139) AhdI (3555) AfeI (3524) BmrI (3515) NmeAIII (3408) BsaHI (3015) XmnI (2955) PsiI (2485) DraIII (2360) BsaAI (2357) BglII (0) NdeI (98) ATG 6xHis NheI (136) BmtI (140) Xpress™ tag EagI - NotI (328) lac UV5 promoter PvuII (548) EcoRI (648) BsmBI (872) PasI - PflMI * (880) BtgI - NcoI (949) BssHII (1142) BstZ17I (1183) BbvCI (1373) AvaI - BsoBI - TspMI - XmaI (1517) BmeT110I (1518) SmaI - SrfI (1519) BmgBI (1553) BstXI (1636) BfuAI - BspMI (1763) PstI (1774) SalI (1776) HindIII (1914) BlpI (1972) EcoO109I (1999) NgoMIV (2252) NaeI (2254) BanII (2286) pEXP1-DEST 4622 bp
BspQI  (4560)
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  (4560)
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  (4447)
1 site
R C A T G Y Y G T A C R
AflIII  (4443)
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  (4443)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PspFI  (4143)
1 site
C C C A G C G G G T C G
BseYI  (4139)
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.
AhdI  (3555)
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.
AfeI  (3524)
1 site
A G C G C T T C G C G A
BmrI  (3515)
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.
NmeAIII  (3408)
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).
BsaHI  (3015)
1 site
G R C G Y C C Y G C R G

BsaHI is typically used at 37°C, but is even more active at 60°C.
XmnI  (2955)
1 site
G A A N N N N T T C C T T N N N N A A G
PsiI  (2485)
1 site
T T A T A A A A T A T T
DraIII  (2360)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
BsaAI  (2357)
1 site
Y A C G T R R T G C A Y
BglII  (0)
1 site
A G A T C T T C T A G A
NdeI  (98)
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.
NheI  (136)
1 site
G C T A G C C G A T C G
BmtI  (140)
1 site
G C T A G C C G A T C G
EagI  (328)
1 site
C G G C C G G C C G G C
NotI  (328)
1 site
G C G G C C G C C G C C G G C G
PvuII  (548)
1 site
C A G C T G G T C G A C
EcoRI  (648)
1 site
G A A T T C C T T A A G
BsmBI  (872)
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.
PasI  (880)
1 site
C C C W G G G G G G W C C C

Sticky ends from different PasI sites may not be compatible.
PflMI  (880)
1 site
C C A N N N N N T G G G G T N N N N N A C C
* Blocked by Dcm methylation.
Sticky ends from different PflMI sites may not be compatible.
BtgI  (949)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
NcoI  (949)
1 site
C C A T G G G G T A C C
BssHII  (1142)
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.
BstZ17I  (1183)
1 site
G T A T A C C A T A T G
BbvCI  (1373)
1 site
C C T C A G C G G A G T C G
AvaI  (1517)
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  (1517)
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.
TspMI  (1517)
1 site
C C C G G G G G G C C C
XmaI  (1517)
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.
BmeT110I  (1518)
1 site
C Y C G R G G R G C Y C
SmaI  (1519)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
SrfI  (1519)
1 site
G C C C G G G C C G G G C C C G
BmgBI  (1553)
1 site
C A C G T C G T G C A G

This recognition sequence is asymmetric, so ligating blunt ends
generated by BmgBI will not always regenerate a BmgBI site.
BstXI  (1636)
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.
BfuAI  (1763)
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  (1763)
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  (1774)
1 site
C T G C A G G A C G T C
SalI  (1776)
1 site
G T C G A C C A G C T G
HindIII  (1914)
1 site
A A G C T T T T C G A A
BlpI  (1972)
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.
EcoO109I  (1999)
1 site
R G G N C C Y Y C C N G G R

Sticky ends from different EcoO109I sites may not be compatible.
NgoMIV  (2252)
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.
NaeI  (2254)
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.
BanII  (2286)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
AmpR
2768 .. 3628  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   2768 .. 2836  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2768 .. 3628  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   2837 .. 3628  =  792 bp
   263 amino acids  =  29.0 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2768 .. 3628  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
CmR
435 .. 1094  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
435 .. 1094  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
3799 .. 4387  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3799 .. 4387  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
2127 .. 2582  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
2127 .. 2582  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
ccdB
1436 .. 1741  =  306 bp
101 amino acids  =  11.7 kDa
Product: CcdB, a bacterial toxin that poisons DNA
gyrase
Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains.
ccdB
1436 .. 1741  =  306 bp
101 amino acids  =  11.7 kDa
Product: CcdB, a bacterial toxin that poisons DNA
gyrase
Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains.
attR1
202 .. 326  =  125 bp
recombination site for the Gateway® LR reaction
attR1
202 .. 326  =  125 bp
recombination site for the Gateway® LR reaction
attR2
1782 .. 1906  =  125 bp
recombination site for the Gateway® LR reaction
attR2
1782 .. 1906  =  125 bp
recombination site for the Gateway® LR reaction
AmpR promoter
2663 .. 2767  =  105 bp
AmpR promoter
2663 .. 2767  =  105 bp
T7 terminator
1983 .. 2030  =  48 bp
transcription terminator for bacteriophage T7 RNA
polymerase
T7 terminator
1983 .. 2030  =  48 bp
transcription terminator for bacteriophage T7 RNA
polymerase
T7 tag (gene 10 leader)
133 .. 165  =  33 bp
11 amino acids  =  1.1 kDa
Product: leader peptide from bacteriophage T7 gene
10
promotes efficient translation in E. coli
T7 tag (gene 10 leader)
133 .. 165  =  33 bp
11 amino acids  =  1.1 kDa
Product: leader peptide from bacteriophage T7 gene
10
promotes efficient translation in E. coli
lac UV5 promoter
351 .. 381  =  31 bp
   Segment 1:  -35  
   351 .. 356  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
351 .. 381  =  31 bp
   Segment 2:  
   357 .. 374  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
351 .. 381  =  31 bp
   Segment 3:  -10  
   375 .. 381  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
351 .. 381  =  31 bp
3 segments
E. coli lac promoter with an "up" mutation
Xpress™ tag
169 .. 192  =  24 bp
8 amino acids  =  998.0 Da
   Segment 1:  
   169 .. 177  =  9 bp
   3 amino acids  =  409.5 Da
Product: Xpress™ epitope tag, including an
enterokinase recognition and cleavage site
Xpress™ tag
169 .. 192  =  24 bp
8 amino acids  =  998.0 Da
   Segment 2:  enterokinase site  
   178 .. 192  =  15 bp
   5 amino acids  =  606.5 Da
Product: Xpress™ epitope tag, including an
enterokinase recognition and cleavage site
Xpress™ tag
169 .. 192  =  24 bp
8 amino acids  =  998.0 Da
2 segments
Product: Xpress™ epitope tag, including an
enterokinase recognition and cleavage site
T7 promoter
20 .. 38  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
20 .. 38  =  19 bp
promoter for bacteriophage T7 RNA polymerase
6xHis
112 .. 129  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
112 .. 129  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
RBS
85 .. 92  =  8 bp
ribosome binding site
RBS
85 .. 92  =  8 bp
ribosome binding site
ATG
100 .. 102  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
100 .. 102  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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