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Plasmid Files

pBAC™-6

Baculovirus transfer plasmid with the gp64 promoter, encoding an N-terminal signal sequence-6xHis-thrombin-S-Tag-enterokinase cassette.

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pBAC-6 Sequence and MappBAC-6.dna
Map and Sequence File   
Sequence Author:  Novagen (EMD Millipore)
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 DraIII (5288) ScaI (4616) NmeAIII (4284) BpmI (4206) BmrI (4176) AlwNI (3659) PspFI (3551) BseYI (3547) PciI (3243) PmeI (3123) SgrAI (2835) BstXI (315) NsiI (332) MluI (547) BbvCI - Bpu10I (564) BclI * (1156) BglII (1161) gp64 promoter SpeI (1323) NcoI (1433) 6xHis SacII (1463) thrombin site PflMI (1524) NheI (1534) BmtI (1538) enterokinase site BseRI - TspMI - XmaI (1566) SmaI - SrfI (1568) StuI * (1586) BamHI (1590) EcoRI (1596) Eco53kI (1604) SacI (1606) HindIII (1609) EagI - NotI (1616) PaeR7I - PspXI - XhoI (1624) 6xHis AvrII (1651) BlpI (1660) SphI (1670) SnaBI (1747) SwaI (1837) ZraI (2026) AatII (2028) BmgBI (2251) AgeI (2595) AleI (2826) pBAC™-6 5580 bp
DraIII  (5288)
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.
ScaI  (4616)
1 site
A G T A C T T C A T G A
NmeAIII  (4284)
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).
BpmI  (4206)
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.
BmrI  (4176)
1 site
A C T G G G ( N ) 4 N T G A C C C ( N ) 4

The 1-base overhangs produced by BmrI may be hard to ligate.
Sticky ends from different BmrI sites may not be compatible.
Unlike most restriction enzymes, BmrI can cleave DNA in the
absence of magnesium.
AlwNI  (3659)
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.
PspFI  (3551)
1 site
C C C A G C G G G T C G
BseYI  (3547)
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  (3243)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PmeI  (3123)
1 site
G T T T A A A C C A A A T T T G
SgrAI  (2835)
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.
BstXI  (315)
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.
NsiI  (332)
1 site
A T G C A T T A C G T A
MluI  (547)
1 site
A C G C G T T G C G C A
BbvCI  (564)
1 site
C C T C A G C G G A G T C G
Bpu10I  (564)
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.
BclI  (1156)
1 site
T G A T C A A C T A G T
* Blocked by Dam methylation.
BclI is typically used at 50-55°C, but is 50% active at 37°C.
BglII  (1161)
1 site
A G A T C T T C T A G A
SpeI  (1323)
1 site
A C T A G T T G A T C A
NcoI  (1433)
1 site
C C A T G G G G T A C C
SacII  (1463)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII
recognition sequence.
PflMI  (1524)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
NheI  (1534)
1 site
G C T A G C C G A T C G
BmtI  (1538)
1 site
G C T A G C C G A T C G
BseRI  (1566)
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.
TspMI  (1566)
1 site
C C C G G G G G G C C C
XmaI  (1566)
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  (1568)
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  (1568)
1 site
G C C C G G G C C G G G C C C G
StuI  (1586)
1 site
A G G C C T T C C G G A
* Blocked by Dcm methylation.
BamHI  (1590)
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.
EcoRI  (1596)
1 site
G A A T T C C T T A A G
Eco53kI  (1604)
1 site
G A G C T C C T C G A G
SacI  (1606)
1 site
G A G C T C C T C G A G
HindIII  (1609)
1 site
A A G C T T T T C G A A
EagI  (1616)
1 site
C G G C C G G C C G G C
NotI  (1616)
1 site
G C G G C C G C C G C C G G C G
PaeR7I  (1624)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (1624)
1 site
V C T C G A G B B G A G C T C V
XhoI  (1624)
1 site
C T C G A G G A G C T C
AvrII  (1651)
1 site
C C T A G G G G A T C C
BlpI  (1660)
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.
SphI  (1670)
1 site
G C A T G C C G T A C G
SnaBI  (1747)
1 site
T A C G T A A T G C A T
SwaI  (1837)
1 site
A T T T A A A T T A A A T T T A

SwaI is typically used at 25°C, but is 50% active at 37°C.
ZraI  (2026)
1 site
G A C G T C C T G C A G
AatII  (2028)
1 site
G A C G T C C T G C A G
BmgBI  (2251)
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.
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.
AleI  (2826)
1 site
C A C N N N N G T G G T G N N N N C A C
baculovirus recombination region (ORF1629)
1684 .. 3041  =  1358 bp
contains part of ORF1629
baculovirus recombination region (ORF1629)
1684 .. 3041  =  1358 bp
contains part of ORF1629
AmpR
4063 .. 4923  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   4063 .. 4854  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4063 .. 4923  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4855 .. 4923  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4063 .. 4923  =  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)
329 .. 1156  =  828 bp
contains ORF603 and part of lef2
baculovirus recombination region (lef2/ORF603)
329 .. 1156  =  828 bp
contains ORF603 and part of lef2
ori
3304 .. 3892  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3304 .. 3892  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
5055 .. 5510  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
5055 .. 5510  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
gp64 promoter
1166 .. 1371  =  206 bp
promoter for the baculovirus gp64 glycoprotein gene
gp64 promoter
1166 .. 1371  =  206 bp
promoter for the baculovirus gp64 glycoprotein gene
AmpR promoter
4924 .. 5028  =  105 bp
AmpR promoter
4924 .. 5028  =  105 bp
MCS
1565 .. 1629  =  65 bp
multiple cloning site
MCS
1565 .. 1629  =  65 bp
multiple cloning site
GP64 signal sequence
1372 .. 1431  =  60 bp
20 amino acids  =  2.0 kDa
Product: signal sequence from the baculovirus
envelope glycoprotein GP64
GP64 signal sequence
1372 .. 1431  =  60 bp
20 amino acids  =  2.0 kDa
Product: signal sequence from the baculovirus
envelope glycoprotein GP64
S-Tag
1489 .. 1533  =  45 bp
15 amino acids  =  1.7 kDa
Product: affinity and epitope tag derived from
pancreatic ribonuclease A
S-Tag
1489 .. 1533  =  45 bp
15 amino acids  =  1.7 kDa
Product: affinity and epitope tag derived from
pancreatic ribonuclease A
6xHis
1441 .. 1458  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
1441 .. 1458  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
thrombin site
1468 .. 1485  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
thrombin site
1468 .. 1485  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
6xHis
1630 .. 1647  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
1630 .. 1647  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
enterokinase site
1549 .. 1563  =  15 bp
5 amino acids  =  606.5 Da
Product: enterokinase recognition and cleavage site
enterokinase site
1549 .. 1563  =  15 bp
5 amino acids  =  606.5 Da
Product: enterokinase recognition and cleavage site
ORF1629
1700 .. 3041  =  1342 bp
446 amino acids  =  49.2 kDa
Product: baculovirus capsid-associated protein
required for viral replication
ORF1629
1700 .. 3041  =  1342 bp
446 amino acids  =  49.2 kDa
Product: baculovirus capsid-associated protein
required for viral replication
ORF603
491 .. 1096  =  606 bp
201 amino acids  =  23.6 kDa
Product: baculovirus ORF603 protein
ORF603
491 .. 1096  =  606 bp
201 amino acids  =  23.6 kDa
Product: baculovirus ORF603 protein
lef2
329 .. 453  =  125 bp
40 amino acids  =  4.5 kDa
Product: baculovirus late expression factor 2
lef2
329 .. 453  =  125 bp
40 amino acids  =  4.5 kDa
Product: baculovirus late expression factor 2
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