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Plasmid Files

pcDNA™-DEST47

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

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pcDNA-DEST47 Sequence and MappcDNA-DEST47.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 SgrDI (7778) PvuI (7227) AhdI (6857) PciI (5964) PfoI (5388) RsrII (5127) PflFI - Tth111I (4729) PluTI (4614) SfoI (4612) NarI (4611) KasI (4610) AvrII (4401) StuI (4400) BseRI (4397) SexAI * (4168) BglII (12) SpeI (249) CMV enhancer SnaBI (590) NotI (1040) EcoRI (1360) BsmBI (1584) PasI - PflMI * (1592) BbvCI (2084) SrfI (2230) BmgBI (2264) BstXI (2347) NheI (2631) BmtI (2635) PaeR7I - PspXI - XhoI (3051) BstBI (3252) BbsI (3564) DraIII (3878) pcDNA™-DEST47 7780 bp
SgrDI  (7778)
1 site
C G T C G A C G G C A G C T G C
PvuI  (7227)
1 site
C G A T C G G C T A G C
AhdI  (6857)
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  (5964)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PfoI  (5388)
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.
RsrII  (5127)
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.
PflFI  (4729)
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  (4729)
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  (4614)
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  (4612)
1 site
G G C G C C C C G C G G
NarI  (4611)
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  (4610)
1 site
G G C G C C C C G C G G
AvrII  (4401)
1 site
C C T A G G G G A T C C
StuI  (4400)
1 site
A G G C C T T C C G G A
BseRI  (4397)
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  (4168)
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
NotI  (1040)
1 site
G C G G C C G C C G C C G G C G
EcoRI  (1360)
1 site
G A A T T C C T T A A G
BsmBI  (1584)
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  (1592)
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  (1592)
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.
BbvCI  (2084)
1 site
C C T C A G C G G A G T C G
SrfI  (2230)
1 site
G C C C G G G C C G G G C C C G
BmgBI  (2264)
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  (2347)
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.
NheI  (2631)
1 site
G C T A G C C G A T C G
BmtI  (2635)
1 site
G C T A G C C G A T C G
PaeR7I  (3051)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (3051)
1 site
V C T C G A G B B G A G C T C V
XhoI  (3051)
1 site
C T C G A G G A G C T C
BstBI  (3252)
1 site
T T C G A A A A G C T T
BbsI  (3564)
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  (3878)
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
6784 .. 7644  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   6784 .. 7575  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
6784 .. 7644  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   7576 .. 7644  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
6784 .. 7644  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
NeoR/KanR
4483 .. 5277  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
4483 .. 5277  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
Cycle 3 GFP
2628 .. 3347  =  720 bp
239 amino acids  =  26.9 kDa
   Segment 1:  
   2628 .. 2630  =  3 bp
   1 amino acid  =  149.2 Da
Product: Cycle 3 GFP (Crameri et al., 1996)
mammalian codon-optimized
Cycle 3 GFP
2628 .. 3347  =  720 bp
239 amino acids  =  26.9 kDa
   Segment 2:  1a  
   2631 .. 2633  =  3 bp
   1 amino acid  =  89.1 Da
Product: Cycle 3 GFP (Crameri et al., 1996)
mammalian codon-optimized
Cycle 3 GFP
2628 .. 3347  =  720 bp
239 amino acids  =  26.9 kDa
   Segment 3:  
   2634 .. 3347  =  714 bp
   237 amino acids  =  26.7 kDa
Product: Cycle 3 GFP (Crameri et al., 1996)
mammalian codon-optimized
Cycle 3 GFP
2628 .. 3347  =  720 bp
239 amino acids  =  26.9 kDa
3 segments
Product: Cycle 3 GFP (Crameri et al., 1996)
mammalian codon-optimized
CmR
1147 .. 1827  =  681 bp
226 amino acids  =  26.6 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
1147 .. 1827  =  681 bp
226 amino acids  =  26.6 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
6025 .. 6613  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
6025 .. 6613  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
3645 .. 4073  =  429 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
3645 .. 4073  =  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
4087 .. 4416  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
4087 .. 4416  =  330 bp
SV40 enhancer and early promoter
ccdB
2147 .. 2452  =  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
2147 .. 2452  =  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
3375 .. 3599  =  225 bp
bovine growth hormone polyadenylation signal
bGH poly(A) signal
3375 .. 3599  =  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
914 .. 1038  =  125 bp
recombination site for the Gateway® LR reaction
attR1
914 .. 1038  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2493 .. 2617  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2493 .. 2617  =  125 bp
recombination site for the Gateway® LR reaction
SV40 poly(A) signal
5453 .. 5574  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
5453 .. 5574  =  122 bp
SV40 polyadenylation signal
AmpR promoter
7645 .. 7749  =  105 bp
AmpR promoter
7645 .. 7749  =  105 bp
lac UV5 promoter
1063 .. 1093  =  31 bp
   Segment 1:  -35  
   1063 .. 1068  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1063 .. 1093  =  31 bp
   Segment 2:  
   1069 .. 1086  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1063 .. 1093  =  31 bp
   Segment 3:  -10  
   1087 .. 1093  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1063 .. 1093  =  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
4267 .. 4402  =  136 bp
SV40 origin of replication
SV40 ori
4267 .. 4402  =  136 bp
SV40 origin of replication
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