pCAMBIA0305.2

Agrobacterium binary vector for plant transformation, with kanamycin-resistance and secreted GUSPlus™ genes.

Sequence Author: Cambia

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NcoI (0) PshAI (9538) PvuII (9070) HindIII (8975) PstI - SbfI (8967) SalI (8957) XbaI (8951) BamHI (8945) SmaI (8942) KpnI - TspMI - XmaI (8940) Acc65I (8936) BanII - SacI (8934) Eco53kI (8932) EcoRI (8924) lac operator AseI (8799) LB T-DNA repeat SacII (8467) PsiI (8339) BpmI (8012) BlpI (7863) BstZ17I (6526) MreI - SgrAI (6324) AgeI (6036) EcoNI (5855) GRP signal sequence BglII (82) SpeI (291) BstBI (631) BsrGI (727) StuI (805) SnaBI (844) AleI (1629) ZraI (1685) AatII (1687) MluI (1771) DraIII (1910) AfeI (2064) 6xHis PmlI (2123) BstEII (2133) AflII (2184) MauBI (2379) PasI (3872) AclI (4466) pCAMBIA0305.2 9737 bp
NcoI  (0)
1 site
C C A T G G G G T A C C
PshAI  (9538)
1 site
G A C N N N N G T C C T G N N N N C A G

PshAI quickly loses activity at 37°C, but can be used at 25°C for long incubations.
PvuII  (9070)
1 site
C A G C T G G T C G A C
HindIII  (8975)
1 site
A A G C T T T T C G A A
PstI  (8967)
1 site
C T G C A G G A C G T C
SbfI  (8967)
1 site
C C T G C A G G G G A C G T C C
SalI  (8957)
1 site
G T C G A C C A G C T G
XbaI  (8951)
1 site
T C T A G A A G A T C T
BamHI  (8945)
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.
SmaI  (8942)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
KpnI  (8940)
1 site
G G T A C C C C A T G G
TspMI  (8940)
1 site
C C C G G G G G G C C C
XmaI  (8940)
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.
Acc65I  (8936)
1 site
G G T A C C C C A T G G
BanII  (8934)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
SacI  (8934)
1 site
G A G C T C C T C G A G
Eco53kI  (8932)
1 site
G A G C T C C T C G A G
EcoRI  (8924)
1 site
G A A T T C C T T A A G
AseI  (8799)
1 site
A T T A A T T A A T T A
SacII  (8467)
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.
PsiI  (8339)
1 site
T T A T A A A A T A T T
BpmI  (8012)
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.
BlpI  (7863)
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.
BstZ17I  (6526)
1 site
G T A T A C C A T A T G
MreI  (6324)
1 site
C G C C G G C G G C G G C C G C
SgrAI  (6324)
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.
AgeI  (6036)
1 site
A C C G G T T G G C C A
EcoNI  (5855)
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.
BglII  (82)
1 site
A G A T C T T C T A G A
SpeI  (291)
1 site
A C T A G T T G A T C A
BstBI  (631)
1 site
T T C G A A A A G C T T
BsrGI  (727)
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.
StuI  (805)
1 site
A G G C C T T C C G G A
SnaBI  (844)
1 site
T A C G T A A T G C A T
AleI  (1629)
1 site
C A C N N N N G T G G T G N N N N C A C
ZraI  (1685)
1 site
G A C G T C C T G C A G
AatII  (1687)
1 site
G A C G T C C T G C A G
MluI  (1771)
1 site
A C G C G T T G C G C A
DraIII  (1910)
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.
AfeI  (2064)
1 site
A G C G C T T C G C G A
PmlI  (2123)
1 site
C A C G T G G T G C A C
BstEII  (2133)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
AflII  (2184)
1 site
C T T A A G G A A T T C
MauBI  (2379)
1 site
C G C G C G C G G C G C G C G C
PasI  (3872)
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.
AclI  (4466)
1 site
A A C G T T T T G C A A
GRP signal sequence
2 .. 82  =  81 bp
27 amino acids  =  2.8 kDa
Product: signal sequence from Oryza sativa glycine-rich cell wall protein
GRP signal sequence
2 .. 82  =  81 bp
27 amino acids  =  2.8 kDa
Product: signal sequence from Oryza sativa glycine-rich cell wall protein
GUSPlus™
83 .. 2099  =  2017 bp
609 amino acids  =  69.6 kDa
2 segments  =  1827 bp
Product: β-glucuronidase
codon-optimized Staphylococcus gusA gene with a catalase intron to ensure expression in plants but not bacteria
GUSPlus™
83 .. 2099  =  2017 bp
609 amino acids  =  69.6 kDa
2 segments  =  1827 bp
   Segment 1:  
   83 .. 91  =  9 bp
   3 amino acids  =  402.5 Da
Product: β-glucuronidase
codon-optimized Staphylococcus gusA gene with a catalase intron to ensure expression in plants but not bacteria
GUSPlus™
83 .. 2099  =  2017 bp
609 amino acids  =  69.6 kDa
2 segments  =  1827 bp
   Segment 2:  
   282 .. 2099  =  1818 bp
   606 amino acids  =  69.2 kDa
Product: β-glucuronidase
codon-optimized Staphylococcus gusA gene with a catalase intron to ensure expression in plants but not bacteria
GUSPlus™
83 .. 2099  =  2017 bp
609 amino acids  =  69.6 kDa
2 segments  =  1827 bp
Product: β-glucuronidase
codon-optimized Staphylococcus gusA gene with a catalase intron to ensure expression in plants but not bacteria
6xHis
2106 .. 2123  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
2106 .. 2123  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
pVS1 RepA
4811 .. 5884  =  1074 bp
357 amino acids  =  39.9 kDa
Product: replication protein from Pseudomonas plasmid pVS1
pVS1 RepA
4811 .. 5884  =  1074 bp
357 amino acids  =  39.9 kDa
Product: replication protein from Pseudomonas plasmid pVS1
KanR
7489 .. 8283  =  795 bp
264 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin
KanR
7489 .. 8283  =  795 bp
264 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin
pVS1 StaA
3753 .. 4382  =  630 bp
209 amino acids  =  22.1 kDa
Product: stability protein from Pseudomonas plasmid pVS1
pVS1 StaA
3753 .. 4382  =  630 bp
209 amino acids  =  22.1 kDa
Product: stability protein from Pseudomonas plasmid pVS1
ori
6814 .. 7402  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
6814 .. 7402  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
CaMV 35S promoter
9377 .. 9722  =  346 bp
strong constitutive promoter from cauliflower mosaic virus
CaMV 35S promoter
9377 .. 9722  =  346 bp
strong constitutive promoter from cauliflower mosaic virus
NOS terminator
2155 .. 2407  =  253 bp
nopaline synthase terminator and poly(A) signal
NOS terminator
2155 .. 2407  =  253 bp
nopaline synthase terminator and poly(A) signal
lacZα
8910 .. 9143  =  234 bp
77 amino acids  =  8.6 kDa
Product: LacZα fragment of β-galactosidase
lacZα
8910 .. 9143  =  234 bp
77 amino acids  =  8.6 kDa
Product: LacZα fragment of β-galactosidase
pVS1 oriV
5950 .. 6144  =  195 bp
origin of replication for the Pseudomonas plasmid pVS1 (Heeb et al., 2000)
pVS1 oriV
5950 .. 6144  =  195 bp
origin of replication for the Pseudomonas plasmid pVS1 (Heeb et al., 2000)
bom
6488 .. 6628  =  141 bp
basis of mobility region from pBR322
bom
6488 .. 6628  =  141 bp
basis of mobility region from pBR322
lac promoter
8836 .. 8866  =  31 bp
3 segments
   Segment 1:  -35  
   8836 .. 8841  =  6 bp
promoter for the E. coli lac operon
lac promoter
8836 .. 8866  =  31 bp
3 segments
   Segment 2:  
   8842 .. 8859  =  18 bp
promoter for the E. coli lac operon
lac promoter
8836 .. 8866  =  31 bp
3 segments
   Segment 3:  -10  
   8860 .. 8866  =  7 bp
promoter for the E. coli lac operon
lac promoter
8836 .. 8866  =  31 bp
3 segments
promoter for the E. coli lac operon
RB T-DNA repeat
2429 .. 2453  =  25 bp
right border repeat from nopaline C58 T-DNA
RB T-DNA repeat
2429 .. 2453  =  25 bp
right border repeat from nopaline C58 T-DNA
LB T-DNA repeat
8708 .. 8732  =  25 bp
left border repeat from nopaline C58 T-DNA
LB T-DNA repeat
8708 .. 8732  =  25 bp
left border repeat from nopaline C58 T-DNA
lac operator
8874 .. 8890  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lac operator
8874 .. 8890  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
cat1 intron
92 .. 281  =  190 bp
castor bean catalase intron, modified
cat1 intron
92 .. 281  =  190 bp
castor bean catalase intron, modified
MCS
8924 .. 8980  =  57 bp
pUC18/19 multiple cloning site
MCS
8924 .. 8980  =  57 bp
pUC18/19 multiple cloning site
M13 rev
8898 .. 8914  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
8898 .. 8914  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
8984 .. 9000  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
8984 .. 9000  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  2768 .. 3454  =  687 bp
ORF:  228 amino acids  =  25.1 kDa
ORF:  4709 .. 5884  =  1176 bp
ORF:  391 amino acids  =  43.9 kDa
ORF:  7622 .. 7852  =  231 bp
ORF:  76 amino acids  =  8.3 kDa
ORF:  423 .. 2129  =  1707 bp
ORF:  568 amino acids  =  64.7 kDa
ORF:  3753 .. 4382  =  630 bp
ORF:  209 amino acids  =  22.1 kDa
ORF:  8910 .. 9143  =  234 bp
ORF:  77 amino acids  =  8.6 kDa
ORF:  9195 .. 9623  =  429 bp
ORF:  142 amino acids  =  15.8 kDa
ORF:  276 .. 2225  =  1950 bp
ORF:  649 amino acids  =  73.4 kDa
ORF:  3219 .. 3491  =  273 bp
ORF:  90 amino acids  =  9.9 kDa
ORF:  1463 .. 1690  =  228 bp
ORF:  75 amino acids  =  7.7 kDa
ORF:  3491 .. 3805  =  315 bp
ORF:  104 amino acids  =  12.0 kDa
ORF:  4679 .. 5251  =  573 bp
ORF:  190 amino acids  =  20.6 kDa
ORF:  8528 .. 8821  =  294 bp
ORF:  97 amino acids  =  10.6 kDa
ORF:  2800 .. 3150  =  351 bp
ORF:  116 amino acids  =  12.6 kDa
ORF:  4657 .. 4914  =  258 bp
ORF:  85 amino acids  =  9.6 kDa
ORF:  7489 .. 8283  =  795 bp
ORF:  264 amino acids  =  31.0 kDa
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