pDNR-Dual

Donor vector for transferring a cloned gene into an acceptor vector, and for tagging the gene, using Cre recombinase in the Creator™ system.

Sequence Author: Clontech (TaKaRa)

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loxP BmtI (4888) NheI (4884) T7 promoter BglII (4856) MluI (4833) PspFI (4478) BseYI (4474) AlwNI (4369) AhdI (3890) BmrI (3850) BsaI (3824) BsrFI (3805) BglI (3772) NmeAIII (3743) AseI (3715) FspI (3667) PvuI (3521) XmnI (3290) SacII (2588) SalI (45) NdeI (61) TspMI - XmaI (66) SmaI (68) EcoRI (71) PstI - SbfI (81) BamHI (82) PaeR7I - XhoI (89) HindIII (95) XbaI (101) BssHII (118) PspOMI * (124) ApaI * (128) splice donor site 6xHN stop AvrII (180) SpeI (200) Eco53kI (314) SacI (316) NcoI (467) PasI (537) PflMI * (543) BsmBI - Esp3I (544) BspEI (772) loxP BfuAI - BspMI (1079) BstZ17I (1380) StuI * (1750) BsgI (2074) PsiI (2149) BsrGI (2189) BsaAI - SnaBI (2258) pDNR-Dual 4938 bp
BmtI  (4888)
1 site
G C T A G C C G A T C G
NheI  (4884)
1 site
G C T A G C C G A T C G
BglII  (4856)
1 site
A G A T C T T C T A G A
MluI  (4833)
1 site
A C G C G T T G C G C A
PspFI  (4478)
1 site
C C C A G C G G G T C G
BseYI  (4474)
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  (4369)
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.
AhdI  (3890)
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.
BmrI  (3850)
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.
BsaI  (3824)
1 site
G G T C T C N C C A G A G N ( N ) 4

Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
BsrFI  (3805)
1 site
R C C G G Y Y G G C C R

Cleavage may be enhanced when more than one copy of the BsrFI recognition sequence is present.
After cleavage, BsrFI can remain bound to DNA and alter its electrophoretic mobility.
BglI  (3772)
1 site
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
NmeAIII  (3743)
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).
AseI  (3715)
1 site
A T T A A T T A A T T A
FspI  (3667)
1 site
T G C G C A A C G C G T
PvuI  (3521)
1 site
C G A T C G G C T A G C
XmnI  (3290)
1 site
G A A N N N N T T C C T T N N N N A A G
SacII  (2588)
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.
SalI  (45)
1 site
G T C G A C C A G C T G
NdeI  (61)
1 site
C A T A T G G T A T A C

Prolonged incubation with NdeI may lead to removal of additional nucleotides.
TspMI  (66)
1 site
C C C G G G G G G C C C
XmaI  (66)
1 site
C C C G G G G G G C C C

Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
SmaI  (68)
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  (71)
1 site
G A A T T C C T T A A G
PstI  (81)
1 site
C T G C A G G A C G T C
SbfI  (81)
1 site
C C T G C A G G G G A C G T C C
BamHI  (82)
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.
PaeR7I  (89)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (89)
1 site
C T C G A G G A G C T C
HindIII  (95)
1 site
A A G C T T T T C G A A
XbaI  (101)
1 site
T C T A G A A G A T C T
BssHII  (118)
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.
PspOMI  (124)
1 site
G G G C C C C C C G G G
* Blocked by Dcm methylation.
ApaI  (128)
1 site
G G G C C C C C C G G G
* Blocked by Dcm methylation.
ApaI can be used between 25°C and 37°C.
AvrII  (180)
1 site
C C T A G G G G A T C C
SpeI  (200)
1 site
A C T A G T T G A T C A
Eco53kI  (314)
1 site
G A G C T C C T C G A G
SacI  (316)
1 site
G A G C T C C T C G A G
NcoI  (467)
1 site
C C A T G G G G T A C C
PasI  (537)
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.
PflMI  (543)
1 site
C C A N N N N N T G G G G T N N N N N A C C
* Blocked by Dcm methylation.
Sticky ends from different PflMI sites may not be compatible.
BsmBI  (544)
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.
BsmBI-v2 is an improved version of BsmBI.
Esp3I  (544)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different Esp3I sites may not be compatible.
BspEI  (772)
1 site
T C C G G A A G G C C T
BfuAI  (1079)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BfuAI recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (1079)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BspMI recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
BstZ17I  (1380)
1 site
G T A T A C C A T A T G
StuI  (1750)
1 site
A G G C C T T C C G G A
* Blocked by Dcm methylation.
BsgI  (2074)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14

Efficient cleavage requires at least two copies of the BsgI recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
PsiI  (2149)
1 site
T T A T A A A A T A T T
BsrGI  (2189)
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.
BsaAI  (2258)
1 site
Y A C G T R R T G C A Y
SnaBI  (2258)
1 site
T A C G T A A T G C A T
SacB
1509 .. 2930  =  1422 bp
473 amino acids  =  53.0 kDa
2 segments
   Segment 1:  signal peptide  
   1509 .. 1595  =  87 bp
   29 amino acids  =  3.0 kDa
Product: secreted levansucrase that renders bacterial growth sensitive to sucrose
negative selection marker
SacB
1509 .. 2930  =  1422 bp
473 amino acids  =  53.0 kDa
2 segments
   Segment 2:  
   1596 .. 2930  =  1335 bp
   444 amino acids  =  50.0 kDa
Product: secreted levansucrase that renders bacterial growth sensitive to sucrose
negative selection marker
SacB
1509 .. 2930  =  1422 bp
473 amino acids  =  53.0 kDa
2 segments
Product: secreted levansucrase that renders bacterial growth sensitive to sucrose
negative selection marker
AmpR
3103 .. 3963  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   3103 .. 3171  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3103 .. 3963  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   3172 .. 3963  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3103 .. 3963  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
CmR
327 .. 986  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
327 .. 986  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
4134 .. 4722  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
4134 .. 4722  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
sacB promoter
1063 .. 1508  =  446 bp
sacB promoter and control region
sacB promoter
1063 .. 1508  =  446 bp
sacB promoter and control region
AmpR promoter
2998 .. 3102  =  105 bp
AmpR promoter
2998 .. 3102  =  105 bp
lambda t0 terminator
212 .. 306  =  95 bp
transcription terminator from phage lambda
lambda t0 terminator
212 .. 306  =  95 bp
transcription terminator from phage lambda
MCS
45 .. 129  =  85 bp
multiple cloning site
MCS
45 .. 129  =  85 bp
multiple cloning site
6xHN
141 .. 176  =  36 bp
12 amino acids  =  1.5 kDa
Product: 6x(His-Asn) affinity tag
6xHN
141 .. 176  =  36 bp
12 amino acids  =  1.5 kDa
Product: 6x(His-Asn) affinity tag
loxP
9 .. 42  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
9 .. 42  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
1015 .. 1048  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
1015 .. 1048  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
T7 promoter
4863 .. 4881  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
4863 .. 4881  =  19 bp
promoter for bacteriophage T7 RNA polymerase
M13 fwd
4839 .. 4855  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
4839 .. 4855  =  17 bp
common sequencing primer, one of multiple similar variants
splice donor site
132 .. 137  =  6 bp
splice donor site
132 .. 137  =  6 bp
stop
177 .. 179  =  3 bp
stop codon
stop
177 .. 179  =  3 bp
stop codon
ORF:  3103 .. 3963  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  1509 .. 2930  =  1422 bp
ORF:  473 amino acids  =  53.0 kDa
ORF:  327 .. 1031  =  705 bp
ORF:  234 amino acids  =  27.3 kDa
ORF:  3567 .. 3833  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  176 .. 409  =  234 bp
ORF:  77 amino acids  =  8.4 kDa
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Download pDNR-Dual.dna file

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