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pFastBac™ Dual

Dual-promoter vector for co-expression of two proteins in insect cells using the Bac-to-Bac® baculovirus system.

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pFastBac Dual Sequence and MappFastBac Dual.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 AvrII (4978) HpaI (4827) MfeI (4814) HindIII (4699) PstI (4690) XbaI (4678) BstBI (4673) NotI (4664) SpeI (4656) SacI (4654) Eco53kI (4652) SalI - SgrDI (4644) StuI (4638) EcoRI (4628) BssHII (4621) RsrII (4613) BamHI (4606) BstZ17I (4458) PacI (4384) BbsI (4325) SmaI (4321) TspMI - XmaI (4319) PaeR7I - XhoI (4311) MCS NcoI (4305) BmtI (4302) NheI (4298) PvuII (4295) NsiI (4291) SphI (4284) KpnI (4278) Acc65I (4274) HSV TK poly(A) signal SnaBI (3982) BsrGI (3510) PflFI - Tth111I (3328) BsmBI (3282) BseRI (3061) EcoRV (2925) SacII (2868) BstXI (2817) NgoMIV (227) NaeI (229) BsaHI (936) PvuI (1107) FspI (1253) BpmI (1407) AlwNI (1955) BseYI (2060) PspFI (2064) BspQI - SapI (2481) MscI (2809) pFastBac™ Dual 5238 bp
AvrII  (4978)
1 site
C C T A G G G G A T C C
HpaI  (4827)
1 site
G T T A A C C A A T T G
MfeI  (4814)
1 site
C A A T T G G T T A A C
HindIII  (4699)
1 site
A A G C T T T T C G A A
PstI  (4690)
1 site
C T G C A G G A C G T C
XbaI  (4678)
1 site
T C T A G A A G A T C T
BstBI  (4673)
1 site
T T C G A A A A G C T T
NotI  (4664)
1 site
G C G G C C G C C G C C G G C G
SpeI  (4656)
1 site
A C T A G T T G A T C A
SacI  (4654)
1 site
G A G C T C C T C G A G
Eco53kI  (4652)
1 site
G A G C T C C T C G A G
SalI  (4644)
1 site
G T C G A C C A G C T G
SgrDI  (4644)
1 site
C G T C G A C G G C A G C T G C
StuI  (4638)
1 site
A G G C C T T C C G G A
EcoRI  (4628)
1 site
G A A T T C C T T A A G
BssHII  (4621)
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.
RsrII  (4613)
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.
BamHI  (4606)
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.
BstZ17I  (4458)
1 site
G T A T A C C A T A T G
PacI  (4384)
1 site
T T A A T T A A A A T T A A T T
BbsI  (4325)
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.
SmaI  (4321)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
TspMI  (4319)
1 site
C C C G G G G G G C C C
XmaI  (4319)
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.
PaeR7I  (4311)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (4311)
1 site
C T C G A G G A G C T C
NcoI  (4305)
1 site
C C A T G G G G T A C C
BmtI  (4302)
1 site
G C T A G C C G A T C G
NheI  (4298)
1 site
G C T A G C C G A T C G
PvuII  (4295)
1 site
C A G C T G G T C G A C
NsiI  (4291)
1 site
A T G C A T T A C G T A
SphI  (4284)
1 site
G C A T G C C G T A C G
KpnI  (4278)
1 site
G G T A C C C C A T G G
Acc65I  (4274)
1 site
G G T A C C C C A T G G
SnaBI  (3982)
1 site
T A C G T A A T G C A T
BsrGI  (3510)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
PflFI  (3328)
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  (3328)
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.
BsmBI  (3282)
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.
BseRI  (3061)
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  (2925)
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.
SacII  (2868)
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.
BstXI  (2817)
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.
NgoMIV  (227)
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.
NaeI  (229)
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.
BsaHI  (936)
1 site
G R C G Y C C Y G C R G

BsaHI is typically used at 37°C, but is even more active at 60°C.
PvuI  (1107)
1 site
C G A T C G G C T A G C
FspI  (1253)
1 site
T G C G C A A C G C G T
BpmI  (1407)
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.
AlwNI  (1955)
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.
BseYI  (2060)
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.
PspFI  (2064)
1 site
C C C A G C G G G T C G
BspQI  (2481)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (2481)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be
mixed before removing an aliquot.
MscI  (2809)
1 site
T G G C C A A C C G G T
AmpR
689 .. 1549  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   689 .. 757  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
689 .. 1549  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   758 .. 1549  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
689 .. 1549  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
1720 .. 2308  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1720 .. 2308  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
GmR
2902 .. 3435  =  534 bp
177 amino acids  =  19.4 kDa
Product: gentamycin acetyltransferase
confers resistance to gentamycin
GmR
2902 .. 3435  =  534 bp
177 amino acids  =  19.4 kDa
Product: gentamycin acetyltransferase
confers resistance to gentamycin
f1 ori
102 .. 557  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
102 .. 557  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
Tn7R
2611 .. 2835  =  225 bp
mini-Tn7 element (right end of the Tn7 transposon)
Tn7R
2611 .. 2835  =  225 bp
mini-Tn7 element (right end of the Tn7 transposon)
Tn7L
4991 .. 5156  =  166 bp
mini-Tn7 element (left end of the Tn7 transposon)
Tn7L
4991 .. 5156  =  166 bp
mini-Tn7 element (left end of the Tn7 transposon)
SV40 poly(A) signal
4828 .. 4962  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
4828 .. 4962  =  135 bp
SV40 polyadenylation signal
p10 promoter
4345 .. 4454  =  110 bp
baculovirus promoter for expression in insect cells
p10 promoter
4345 .. 4454  =  110 bp
baculovirus promoter for expression in insect cells
AmpR promoter
584 .. 688  =  105 bp
AmpR promoter
584 .. 688  =  105 bp
MCS
4606 .. 4704  =  99 bp
multiple cloning site
MCS
4606 .. 4704  =  99 bp
multiple cloning site
polyhedrin promoter
4478 .. 4569  =  92 bp
promoter for the baculovirus polyhedrin gene
polyhedrin promoter
4478 .. 4569  =  92 bp
promoter for the baculovirus polyhedrin gene
MCS
4274 .. 4337  =  64 bp
multiple cloning site
MCS
4274 .. 4337  =  64 bp
multiple cloning site
HSV TK poly(A) signal
4161 .. 4209  =  49 bp
herpesvirus thymidine kinase polyadenylation signal
HSV TK poly(A) signal
4161 .. 4209  =  49 bp
herpesvirus thymidine kinase polyadenylation signal
Pc promoter
3624 .. 3652  =  29 bp
   Segment 3:  -10  
   3624 .. 3629  =  6 bp
class 1 integron promoter
Pc promoter
3624 .. 3652  =  29 bp
   Segment 2:  
   3630 .. 3646  =  17 bp
class 1 integron promoter
Pc promoter
3624 .. 3652  =  29 bp
   Segment 1:  -35  
   3647 .. 3652  =  6 bp
class 1 integron promoter
Pc promoter
3624 .. 3652  =  29 bp
3 segments
class 1 integron promoter
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