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Plasmid Files

pAcG2T

Baculovirus transfer vector for expressing N-terminally GST-tagged proteins with a thrombin cleavage site, for use with the BD BaculoGold™ system.

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pAcG2T Sequence and MappAcG2T.dna
Map and Sequence File   
Sequence Author:  BD Biosciences
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 PspFI (7870) BseYI (7866) AlwNI (7761) BpmI (7213) NmeAIII (7135) PfoI (6243) DraIII (5886) BanII (5812) EagI (5183) FspAI (4506) SgrDI (4353) SphI (234) BstXI (1255) BbvCI - Bpu10I (1504) BseRI (1620) EcoRV (2099) EcoNI (2216) BamHI (2878) TspMI - XmaI (2883) SmaI (2885) EcoRI (2888) SnaBI (2986) AgeI (3834) SgrAI (4074) pAcG2T 8530 bp
PspFI  (7870)
1 site
C C C A G C G G G T C G
BseYI  (7866)
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.
AlwNI  (7761)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
BpmI  (7213)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI
recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its
electrophoretic mobility.
BpmI quickly loses activity at 37°C.
NmeAIII  (7135)
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).
PfoI  (6243)
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.
DraIII  (5886)
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.
BanII  (5812)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
EagI  (5183)
1 site
C G G C C G G C C G G C
FspAI  (4506)
1 site
R T G C G C A Y Y A C G C G T R
SgrDI  (4353)
1 site
C G T C G A C G G C A G C T G C
SphI  (234)
1 site
G C A T G C C G T A C G
BstXI  (1255)
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.
BbvCI  (1504)
1 site
C C T C A G C G G A G T C G
Bpu10I  (1504)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
This recognition sequence is asymmetric, so ligating sticky ends
generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
BseRI  (1620)
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.
EcoRV  (2099)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to
delete a base after cleavage.
EcoNI  (2216)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
BamHI  (2878)
1 site
G G A T C C C C T A G G

After cleavage, BamHI-HF™ (but not the original BamHI) can
remain bound to DNA and alter its electrophoretic mobility.
TspMI  (2883)
1 site
C C C G G G G G G C C C
XmaI  (2883)
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  (2885)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
EcoRI  (2888)
1 site
G A A T T C C T T A A G
SnaBI  (2986)
1 site
T A C G T A A T G C A T
AgeI  (3834)
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.
SgrAI  (4074)
1 site
C R C C G G Y G G Y G G C C R C

Efficient cleavage requires at least two copies of the SgrAI
recognition sequence.
baculovirus recombination region (ORF1629)
2923 .. 4280  =  1358 bp
contains part of ORF1629
baculovirus recombination region (ORF1629)
2923 .. 4280  =  1358 bp
contains part of ORF1629
AmpR
6495 .. 7355  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   6495 .. 6563  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
6495 .. 7355  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   6564 .. 7355  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
6495 .. 7355  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
baculovirus recombination region (lef2/ORF603)
1269 .. 2096  =  828 bp
contains ORF603 and part of lef2
baculovirus recombination region (lef2/ORF603)
1269 .. 2096  =  828 bp
contains ORF603 and part of lef2
GST
2206 .. 2859  =  654 bp
218 amino acids  =  25.5 kDa
Product: glutathione S-transferase from
Schistosoma japonicum
GST
2206 .. 2859  =  654 bp
218 amino acids  =  25.5 kDa
Product: glutathione S-transferase from
Schistosoma japonicum
ori
7526 .. 8114  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
7526 .. 8114  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
5653 .. 6108  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
5653 .. 6108  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
AmpR promoter
6390 .. 6494  =  105 bp
AmpR promoter
6390 .. 6494  =  105 bp
polyhedrin promoter
2100 .. 2191  =  92 bp
promoter for the baculovirus polyhedrin gene
polyhedrin promoter
2100 .. 2191  =  92 bp
promoter for the baculovirus polyhedrin gene
thrombin site
2866 .. 2883  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
thrombin site
2866 .. 2883  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
ORF1629
2939 .. 4280  =  1342 bp
446 amino acids  =  49.2 kDa
Product: baculovirus capsid-associated protein
required for viral replication
ORF1629
2939 .. 4280  =  1342 bp
446 amino acids  =  49.2 kDa
Product: baculovirus capsid-associated protein
required for viral replication
ORF603
1431 .. 2036  =  606 bp
201 amino acids  =  23.6 kDa
Product: baculovirus ORF603 protein
ORF603
1431 .. 2036  =  606 bp
201 amino acids  =  23.6 kDa
Product: baculovirus ORF603 protein
lef2
1269 .. 1393  =  125 bp
40 amino acids  =  4.5 kDa
Product: baculovirus late expression factor 2
lef2
1269 .. 1393  =  125 bp
40 amino acids  =  4.5 kDa
Product: baculovirus late expression factor 2
MCS
2878 .. 2893  =  16 bp
multiple cloning site
MCS
2878 .. 2893  =  16 bp
multiple cloning site
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