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Plasmid Files

pBAC4x-1

Baculovirus transfer plasmid with two polyhedrin and two p10 promoters, for the simultaneous expression of up to four proteins.

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pBAC4x-1 Sequence and MappBAC4x-1.dna
Map and Sequence File   
Sequence Author:  Novagen (EMD Millipore)
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 DraIII (5625) ScaI (4953) NmeAIII (4621) BpmI (4543) BmrI (4513) AlwNI (3996) PspFI (3888) BseYI (3884) PciI (3580) PmeI (3460) SgrAI (3172) AleI (3163) BsmI (85) BstXI (315) BbvCI - Bpu10I (564) poly(A) signal Bsu36I (1223) MCS EcoRI (1237) BglII (1243) PpuMI (1357) StuI (1495) MCS XbaI (1498) polyhedrin promoter PspOMI (1621) ApaI (1625) TspMI - XmaI (1744) SmaI (1746) MCS SpeI (1750) BamHI (1937) HindIII (1946) MCS EagI - NotI (1953) PaeR7I - PspXI - XhoI (1961) 6xHis AvrII - StyI (1988) BlpI (1997) SphI (2007) SnaBI (2084) SwaI (2174) ZraI (2363) AatII (2365) BmgBI (2588) AgeI (2932) pBAC4x-1 5917 bp
DraIII  (5625)
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  (4953)
1 site
A G T A C T T C A T G A
NmeAIII  (4621)
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  (4543)
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  (4513)
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  (3996)
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  (3888)
1 site
C C C A G C G G G T C G
BseYI  (3884)
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  (3580)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PmeI  (3460)
1 site
G T T T A A A C C A A A T T T G
SgrAI  (3172)
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.
AleI  (3163)
1 site
C A C N N N N G T G G T G N N N N C A C
BsmI  (85)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
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.
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.
Bsu36I  (1223)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
EcoRI  (1237)
1 site
G A A T T C C T T A A G
BglII  (1243)
1 site
A G A T C T T C T A G A
PpuMI  (1357)
1 site
R G G W C C Y Y C C W G G R

Sticky ends from different PpuMI sites may not be compatible.
StuI  (1495)
1 site
A G G C C T T C C G G A
XbaI  (1498)
1 site
T C T A G A A G A T C T
PspOMI  (1621)
1 site
G G G C C C C C C G G G
ApaI  (1625)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
TspMI  (1744)
1 site
C C C G G G G G G C C C
XmaI  (1744)
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  (1746)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
SpeI  (1750)
1 site
A C T A G T T G A T C A
BamHI  (1937)
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.
HindIII  (1946)
1 site
A A G C T T T T C G A A
EagI  (1953)
1 site
C G G C C G G C C G G C
NotI  (1953)
1 site
G C G G C C G C C G C C G G C G
PaeR7I  (1961)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (1961)
1 site
V C T C G A G B B G A G C T C V
XhoI  (1961)
1 site
C T C G A G G A G C T C
AvrII  (1988)
1 site
C C T A G G G G A T C C
StyI  (1988)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
BlpI  (1997)
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  (2007)
1 site
G C A T G C C G T A C G
SnaBI  (2084)
1 site
T A C G T A A T G C A T
SwaI  (2174)
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  (2363)
1 site
G A C G T C C T G C A G
AatII  (2365)
1 site
G A C G T C C T G C A G
BmgBI  (2588)
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  (2932)
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.
baculovirus recombination region (ORF1629)
2021 .. 3378  =  1358 bp
contains part of ORF1629
baculovirus recombination region (ORF1629)
2021 .. 3378  =  1358 bp
contains part of ORF1629
AmpR
4400 .. 5260  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   4400 .. 5191  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4400 .. 5260  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   5192 .. 5260  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4400 .. 5260  =  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
3641 .. 4229  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3641 .. 4229  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
5392 .. 5847  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
5392 .. 5847  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
p10 promoter
1251 .. 1360  =  110 bp
baculovirus promoter for expression in insect cells
p10 promoter
1251 .. 1360  =  110 bp
baculovirus promoter for expression in insect cells
p10 promoter
1627 .. 1736  =  110 bp
baculovirus promoter for expression in insect cells
p10 promoter
1627 .. 1736  =  110 bp
baculovirus promoter for expression in insect cells
AmpR promoter
5261 .. 5365  =  105 bp
AmpR promoter
5261 .. 5365  =  105 bp
polyhedrin promoter
1511 .. 1602  =  92 bp
promoter for the baculovirus polyhedrin gene
polyhedrin promoter
1511 .. 1602  =  92 bp
promoter for the baculovirus polyhedrin gene
polyhedrin promoter
1842 .. 1933  =  92 bp
promoter for the baculovirus polyhedrin gene
polyhedrin promoter
1842 .. 1933  =  92 bp
promoter for the baculovirus polyhedrin gene
poly(A) signal
1171 .. 1219  =  49 bp
synthetic polyadenylation signal
poly(A) signal
1171 .. 1219  =  49 bp
synthetic polyadenylation signal
poly(A) signal
1756 .. 1804  =  49 bp
synthetic polyadenylation signal
poly(A) signal
1756 .. 1804  =  49 bp
synthetic polyadenylation signal
MCS
1937 .. 1966  =  30 bp
multiple cloning site
MCS
1937 .. 1966  =  30 bp
multiple cloning site
MCS
1222 .. 1248  =  27 bp
multiple cloning site
MCS
1222 .. 1248  =  27 bp
multiple cloning site
6xHis
1967 .. 1984  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
1967 .. 1984  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
MCS
1744 .. 1755  =  12 bp
multiple cloning site
MCS
1744 .. 1755  =  12 bp
multiple cloning site
MCS
1493 .. 1503  =  11 bp
multiple cloning site
MCS
1493 .. 1503  =  11 bp
multiple cloning site
ORF1629
2037 .. 3378  =  1342 bp
446 amino acids  =  49.2 kDa
Product: baculovirus capsid-associated protein
required for viral replication
ORF1629
2037 .. 3378  =  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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