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Plasmid Files

pANT7_nHA

Gateway® destination vector for in vitro transcription and translation of a protein with an N-terminal HA tag.

 
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pANT7_nHA.dna
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PspOMI (55) NaeI (5027) NgoMIV (5025) PsiI (4796) PfoI (4507) AatII (4396) ZraI (4394) NmeAIII (3622) AhdI (3474) PspFI (2889) BseYI (2885) ApaI (59) AvrII (93) PmlI (258) AarI (281) Acc65I (383) KpnI (387) BspEI (1009) EcoRI (1013) PasI (1245) NcoI (1314) BglII (1460) BssHII (1507) BstZ17I (1548) BbvCI (1738) TspMI - XmaI (1882) SmaI - SrfI (1884) BstXI (2001) PstI (2139) SalI (2141) PaeR7I - PspXI - XhoI (2295) PmeI (2370) SpeI (2375) BlpI (2384) pANT7_nHA 5339 bp
PspOMI  (55)
1 site
G G G C C C C C C G G G
NaeI  (5027)
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.
NgoMIV  (5025)
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.
PsiI  (4796)
1 site
T T A T A A A A T A T T
PfoI  (4507)
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.
AatII  (4396)
1 site
G A C G T C C T G C A G
ZraI  (4394)
1 site
G A C G T C C T G C A G
NmeAIII  (3622)
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  (3474)
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  (2889)
1 site
C C C A G C G G G T C G
BseYI  (2885)
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.
ApaI  (59)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
AvrII  (93)
1 site
C C T A G G G G A T C C
PmlI  (258)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
AarI  (281)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the AarI recognition sequence.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its electrophoretic mobility.
Acc65I  (383)
1 site
G G T A C C C C A T G G
KpnI  (387)
1 site
G G T A C C C C A T G G
BspEI  (1009)
1 site
T C C G G A A G G C C T
EcoRI  (1013)
1 site
G A A T T C C T T A A G
PasI  (1245)
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.
NcoI  (1314)
1 site
C C A T G G G G T A C C
BglII  (1460)
1 site
A G A T C T T C T A G A
BssHII  (1507)
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  (1548)
1 site
G T A T A C C A T A T G
BbvCI  (1738)
1 site
C C T C A G C G G A G T C G
TspMI  (1882)
1 site
C C C G G G G G G C C C
XmaI  (1882)
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.
SmaI  (1884)
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  (1884)
1 site
G C C C G G G C C G G G C C C G
BstXI  (2001)
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.
PstI  (2139)
1 site
C T G C A G G A C G T C
SalI  (2141)
1 site
G T C G A C C A G C T G
PaeR7I  (2295)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (2295)
1 site
V C T C G A G B B G A G C T C V
XhoI  (2295)
1 site
C T C G A G G A G C T C
PmeI  (2370)
1 site
G T T T A A A C C A A A T T T G
SpeI  (2375)
1 site
A C T A G T T G A T C A
BlpI  (2384)
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.
AmpR
3401 .. 4261  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   3401 .. 4192  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3401 .. 4261  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4193 .. 4261  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3401 .. 4261  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
CmR
800 .. 1459  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
800 .. 1459  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
2642 .. 3230  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2642 .. 3230  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
IRES
17 .. 515  =  499 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
IRES
17 .. 515  =  499 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
f1 ori
4700 .. 5155  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
4700 .. 5155  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
ccdB
1801 .. 2106  =  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
1801 .. 2106  =  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
567 .. 691  =  125 bp
recombination site for the Gateway® LR reaction
attR1
567 .. 691  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2147 .. 2271  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2147 .. 2271  =  125 bp
recombination site for the Gateway® LR reaction
AmpR promoter
4262 .. 4366  =  105 bp
AmpR promoter
4262 .. 4366  =  105 bp
T7 terminator
2395 .. 2442  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
2395 .. 2442  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
lac UV5 promoter
716 .. 746  =  31 bp
   Segment 1:  -35  
   716 .. 721  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
716 .. 746  =  31 bp
   Segment 2:  
   722 .. 739  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
716 .. 746  =  31 bp