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Plasmid Files

pcDNA™-DEST53

Gateway® destination vector for expressing N-terminally GFP-tagged proteins in mammalian cells.

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pcDNA-DEST53 Sequence and MappcDNA-DEST53.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 SgrDI (7765) PvuI (7214) AhdI (6844) PciI (5951) PfoI (5375) PflFI - Tth111I (4718) PluTI (4603) SfoI (4601) NarI (4600) KasI (4599) AvrII (4390) StuI (4389) BseRI (4386) SexAI * (4157) BglII (12) SpeI (249) CMV enhancer SnaBI (590) HindIII (1028) PaeR7I - PspXI - XhoI (1328) NotI (1769) EcoRI (2089) BsmBI (2313) PasI - PflMI * (2321) AgeI (2595) BbvCI (2793) SrfI (2939) BmgBI (2973) BstXI (3056) PacI (3336) BbsI (3553) DraIII (3867) pcDNA™-DEST53 7767 bp
SgrDI  (7765)
1 site
C G T C G A C G G C A G C T G C
PvuI  (7214)
1 site
C G A T C G G C T A G C
AhdI  (6844)
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  (5951)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PfoI  (5375)
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.
PflFI  (4718)
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  (4718)
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.
PluTI  (4603)
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.
SfoI  (4601)
1 site
G G C G C C C C G C G G
NarI  (4600)
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.
KasI  (4599)
1 site
G G C G C C C C G C G G
AvrII  (4390)
1 site
C C T A G G G G A T C C
StuI  (4389)
1 site
A G G C C T T C C G G A
BseRI  (4386)
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.
SexAI  (4157)
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.
BglII  (12)
1 site
A G A T C T T C T A G A
SpeI  (249)
1 site
A C T A G T T G A T C A
SnaBI  (590)
1 site
T A C G T A A T G C A T
HindIII  (1028)
1 site
A A G C T T T T C G A A
PaeR7I  (1328)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (1328)
1 site
V C T C G A G B B G A G C T C V
XhoI  (1328)
1 site
C T C G A G G A G C T C
NotI  (1769)
1 site
G C G G C C G C C G C C G G C G
EcoRI  (2089)
1 site
G A A T T C C T T A A G
BsmBI  (2313)
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  (2321)
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  (2321)
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.
AgeI  (2595)
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.
BbvCI  (2793)
1 site
C C T C A G C G G A G T C G
SrfI  (2939)
1 site
G C C C G G G C C G G G C C C G
BmgBI  (2973)
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  (3056)
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.
PacI  (3336)
1 site
T T A A T T A A A A T T A A T T
BbsI  (3553)
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.
DraIII  (3867)
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.
AmpR
6771 .. 7631  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   6771 .. 7562  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
6771 .. 7631  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   7563 .. 7631  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
6771 .. 7631  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
NeoR/KanR
4472 .. 5266  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
4472 .. 5266  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
Cycle 3 GFP
905 .. 1621  =  717 bp
239 amino acids  =  26.9 kDa
   Segment 1:  
   905 .. 907  =  3 bp
   1 amino acid  =  149.2 Da
Product: Cycle 3 GFP (Crameri et al., 1996)
mammalian codon-optimized
Cycle 3 GFP
905 .. 1621  =  717 bp
239 amino acids  =  26.9 kDa
   Segment 2:  1a  
   908 .. 910  =  3 bp
   1 amino acid  =  89.1 Da
Product: Cycle 3 GFP (Crameri et al., 1996)
mammalian codon-optimized
Cycle 3 GFP
905 .. 1621  =  717 bp
239 amino acids  =  26.9 kDa
   Segment 3:  
   911 .. 1621  =  711 bp
   237 amino acids  =  26.7 kDa
Product: Cycle 3 GFP (Crameri et al., 1996)
mammalian codon-optimized
Cycle 3 GFP
905 .. 1621  =  717 bp
239 amino acids  =  26.9 kDa
3 segments
Product: Cycle 3 GFP (Crameri et al., 1996)
mammalian codon-optimized
CmR
1876 .. 2535  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
1876 .. 2535  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
6012 .. 6600  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
6012 .. 6600  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
3634 .. 4062  =  429 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
3634 .. 4062  =  429 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
CMV enhancer
235 .. 614  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
235 .. 614  =  380 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
4076 .. 4405  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
4076 .. 4405  =  330 bp
SV40 enhancer and early promoter
ccdB
2856 .. 3161  =  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
2856 .. 3161  =  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.
bGH poly(A) signal
3364 .. 3588  =  225 bp
bovine growth hormone polyadenylation signal
bGH poly(A) signal
3364 .. 3588  =  225 bp
bovine growth hormone polyadenylation signal
CMV promoter
615 .. 818  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
615 .. 818  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
attR1
1643 .. 1767  =  125 bp
recombination site for the Gateway® LR reaction
attR1
1643 .. 1767  =  125 bp
recombination site for the Gateway® LR reaction
attR2
3202 .. 3326  =  125 bp
recombination site for the Gateway® LR reaction
attR2
3202 .. 3326  =  125 bp
recombination site for the Gateway® LR reaction
SV40 poly(A) signal
5440 .. 5561  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
5440 .. 5561  =  122 bp
SV40 polyadenylation signal
AmpR promoter
7632 .. 7736  =  105 bp
AmpR promoter
7632 .. 7736  =  105 bp
lac UV5 promoter
1792 .. 1822  =  31 bp
   Segment 1:  -35  
   1792 .. 1797  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1792 .. 1822  =  31 bp
   Segment 2:  
   1798 .. 1815  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1792 .. 1822  =  31 bp
   Segment 3:  -10  
   1816 .. 1822  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1792 .. 1822  =  31 bp
3 segments
E. coli lac promoter with an "up" mutation
T7 promoter
863 .. 881  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
863 .. 881  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SV40 ori
4256 .. 4391  =  136 bp
SV40 origin of replication
SV40 ori
4256 .. 4391  =  136 bp
SV40 origin of replication
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