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Plasmid Files

pMT/BioEase™-DEST

Gateway® destination vector for inducible high-level expression of biotinylated fusion proteins in Drosophila cells.

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pMT BioEase-DEST Sequence and MappMT BioEase-DEST.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 AatII (5311) ZraI (5309) NmeAIII (4537) AhdI (4389) PspFI (3804) BseYI (3800) PciI (3496) BspQI - SapI (3380) SbfI (3129) MfeI (3020) HpaI (3011) PsiI (2991) ApaI (2837) PspOMI (2833) XbaI * (2827) BglII (2824) PacI (2821) PfoI (46) BstAPI (185) BsgI (640) AflII (852) ATG AleI (879) BstEII (881) NotI (1254) BspEI (1570) EcoRI (1574) PasI (1806) MluI (2021) BssHII (2068) NruI (2076) AgeI (2080) BstZ17I (2088) BbvCI (2278) AvaI - BsoBI - TspMI - XmaI (2422) BmeT110I (2423) SmaI - SrfI (2424) pMT/BioEase™-DEST 5375 bp
AatII  (5311)
1 site
G A C G T C C T G C A G
ZraI  (5309)
1 site
G A C G T C C T G C A G
NmeAIII  (4537)
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).
AhdI  (4389)
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.
PspFI  (3804)
1 site
C C C A G C G G G T C G
BseYI  (3800)
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.
PciI  (3496)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (3380)
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  (3380)
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.
SbfI  (3129)
1 site
C C T G C A G G G G A C G T C C
MfeI  (3020)
1 site
C A A T T G G T T A A C
HpaI  (3011)
1 site
G T T A A C C A A T T G
PsiI  (2991)
1 site
T T A T A A A A T A T T
ApaI  (2837)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PspOMI  (2833)
1 site
G G G C C C C C C G G G
XbaI  (2827)
1 site
T C T A G A A G A T C T
* Blocked by Dam methylation.
BglII  (2824)
1 site
A G A T C T T C T A G A
PacI  (2821)
1 site
T T A A T T A A A A T T A A T T
PfoI  (46)
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.
BstAPI  (185)
1 site
G C A N N N N N T G C C G T N N N N N A C G

Sticky ends from different BstAPI sites may not be compatible.
BsgI  (640)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14

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).
AflII  (852)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
AleI  (879)
1 site
C A C N N N N G T G G T G N N N N C A C
BstEII  (881)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
NotI  (1254)
1 site
G C G G C C G C C G C C G G C G
BspEI  (1570)
1 site
T C C G G A A G G C C T
EcoRI  (1574)
1 site
G A A T T C C T T A A G
PasI  (1806)
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.
MluI  (2021)
1 site
A C G C G T T G C G C A
BssHII  (2068)
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.
NruI  (2076)
1 site
T C G C G A A G C G C T
AgeI  (2080)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
BstZ17I  (2088)
1 site
G T A T A C C A T A T G
BbvCI  (2278)
1 site
C C T C A G C G G A G T C G
AvaI  (2422)
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  (2422)
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  (2422)
1 site
C C C G G G G G G C C C
XmaI  (2422)
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  (2423)
1 site
C Y C G R G G R G C Y C
SmaI  (2424)
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  (2424)
1 site
G C C C G G G C C G G G C C C G
AmpR
4316 .. 5176  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   4316 .. 5107  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4316 .. 5176  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   5108 .. 5176  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4316 .. 5176  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
CmR
1361 .. 2020  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
1361 .. 2020  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
3557 .. 4145  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3557 .. 4145  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
MT promoter
407 .. 833  =  427 bp
Drosophila metallothionein promoter
MT promoter
407 .. 833  =  427 bp
Drosophila metallothionein promoter
ccdB
2341 .. 2646  =  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
2341 .. 2646  =  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.
BioEase™ tag
867 .. 1082  =  216 bp
72 amino acids  =  7.1 kDa
   Segment 1:  
   867 .. 977  =  111 bp
   37 amino acids  =  3.6 kDa
Product: biotinylated sequence derived from the
C-terminus of the α-subunit of Klebsiella
pneumoniae
oxaloacetate decarboxylase
contains a unique biotinylation site
BioEase™ tag
867 .. 1082  =  216 bp
72 amino acids  =  7.1 kDa
   Segment 2:  site  
   978 .. 980  =  3 bp
   1 amino acid  =  146.2 Da
Product: biotinylated sequence derived from the
C-terminus of the α-subunit of Klebsiella
pneumoniae
oxaloacetate decarboxylase
contains a unique biotinylation site
BioEase™ tag
867 .. 1082  =  216 bp
72 amino acids  =  7.1 kDa
   Segment 3:  
   981 .. 1082  =  102 bp
   34 amino acids  =  3.4 kDa
Product: biotinylated sequence derived from the
C-terminus of the α-subunit of Klebsiella
pneumoniae
oxaloacetate decarboxylase
contains a unique biotinylation site
BioEase™ tag
867 .. 1082  =  216 bp
72 amino acids  =  7.1 kDa
3 segments
Product: biotinylated sequence derived from the
C-terminus of the α-subunit of Klebsiella
pneumoniae
oxaloacetate decarboxylase
contains a unique biotinylation site
SV40 poly(A) signal
2877 .. 3011  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2877 .. 3011  =  135 bp
SV40 polyadenylation signal
attR1
1128 .. 1252  =  125 bp
recombination site for the Gateway® LR reaction
attR1
1128 .. 1252  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2687 .. 2811  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2687 .. 2811  =  125 bp
recombination site for the Gateway® LR reaction
AmpR promoter
5177 .. 5281  =  105 bp
AmpR promoter
5177 .. 5281  =  105 bp
lac UV5 promoter
1277 .. 1307  =  31 bp
   Segment 1:  -35  
   1277 .. 1282  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1277 .. 1307  =  31 bp
   Segment 2:  
   1283 .. 1300  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1277 .. 1307  =  31 bp
   Segment 3:  -10  
   1301 .. 1307  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1277 .. 1307  =  31 bp
3 segments
E. coli lac promoter with an "up" mutation
Xpress™ tag
1095 .. 1118  =  24 bp
8 amino acids  =  998.0 Da
   Segment 1:  
   1095 .. 1103  =  9 bp
   3 amino acids  =  409.5 Da
Product: Xpress™ epitope tag, including an
enterokinase recognition and cleavage site
Xpress™ tag
1095 .. 1118  =  24 bp
8 amino acids  =  998.0 Da
   Segment 2:  enterokinase site  
   1104 .. 1118  =  15 bp
   5 amino acids  =  606.5 Da
Product: Xpress™ epitope tag, including an
enterokinase recognition and cleavage site
Xpress™ tag
1095 .. 1118  =  24 bp
8 amino acids  =  998.0 Da
2 segments
Product: Xpress™ epitope tag, including an
enterokinase recognition and cleavage site
ATG
864 .. 866  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
864 .. 866  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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