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Plasmid Files

pFGC5941

Agrobacterium binary vector for expressing dsRNA, with two multiple cloning sites separated by an intron.

 
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 LB T-DNA repeat BclI * (11,169) EcoRV (10,289) BstZ17I (9225) PluTI (9030) SfoI (9028) NarI (9027) KasI (9026) EcoNI (8554) BsiWI (8125) Acc65I (407) KpnI (411) FspAI (521) AarI (585) SalI (656) AfeI (758) EcoRI (1317) MluI (1536) StuI (1867) PshAI (2466) PaeR7I - PspXI - XhoI (2653) TMV Ω NcoI (2734) AscI (2741) SwaI (2760) AflII (3659) PmlI (3675) BamHI (4122) AvrII (4126) XbaI (4136) PacI (4149) TspMI - XmaI (4155) SmaI (4157) MfeI (4465) BstEII (4737) AhdI (4867) SbfI (4892) PvuI (5026) PmeI (5113) pFGC5941 11,406 bp
BclI  (11,169)
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.
EcoRV  (10,289)
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.
BstZ17I  (9225)
1 site
G T A T A C C A T A T G
PluTI  (9030)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI
recognition sequence.
SfoI  (9028)
1 site
G G C G C C C C G C G G
NarI  (9027)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI
recognition sequence.
KasI  (9026)
1 site
G G C G C C C C G C G G
EcoNI  (8554)
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.
BsiWI  (8125)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
Acc65I  (407)
1 site
G G T A C C C C A T G G
KpnI  (411)
1 site
G G T A C C C C A T G G
FspAI  (521)
1 site
R T G C G C A Y Y A C G C G T R
AarI  (585)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the AarI
recognition sequence.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its
electrophoretic mobility.
SalI  (656)
1 site
G T C G A C C A G C T G
AfeI  (758)
1 site
A G C G C T T C G C G A
EcoRI  (1317)
1 site
G A A T T C C T T A A G
MluI  (1536)
1 site
A C G C G T T G C G C A
StuI  (1867)
1 site
A G G C C T T C C G G A
PshAI  (2466)
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.
PaeR7I  (2653)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (2653)
1 site
V C T C G A G B B G A G C T C V
XhoI  (2653)
1 site
C T C G A G G A G C T C
NcoI  (2734)
1 site
C C A T G G G G T A C C
AscI  (2741)
1 site
G G C G C G C C C C G C G C G G
SwaI  (2760)
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.
AflII  (3659)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
PmlI  (3675)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
BamHI  (4122)
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.
AvrII  (4126)
1 site
C C T A G G G G A T C C
XbaI  (4136)
1 site
T C T A G A A G A T C T
PacI  (4149)
1 site
T T A A T T A A A A T T A A T T
TspMI  (4155)
1 site
C C C G G G G G G C C C
XmaI  (4155)
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  (4157)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
MfeI  (4465)
1 site
C A A T T G G T T A A C
BstEII  (4737)
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.
AhdI  (4867)
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.
SbfI  (4892)
1 site
C C T G C A G G G G A C G T C C
PvuI  (5026)
1 site
C G A T C G G C T A G C
PmeI  (5113)
1 site
G T T T A A A C C A A A T T T G
chsA intron
2767 .. 4115  =  1349 bp
chalcone synthase A intron from Petunia hybrida
chsA intron
2767 .. 4115  =  1349 bp
chalcone synthase A intron from Petunia hybrida
pVS1 RepA
7510 .. 8583  =  1074 bp
357 amino acids  =  39.9 kDa
Product: replication protein from Pseudomonas
plasmid pVS1
pVS1 RepA
7510 .. 8583  =  1074 bp
357 amino acids  =  39.9 kDa
Product: replication protein from Pseudomonas
plasmid pVS1
KanR
10,188 .. 10,982  =  795 bp
264 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin
KanR
10,188 .. 10,982  =  795 bp
264 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin
OCS terminator
4177 .. 4884  =  708 bp
octopine synthase terminator
OCS terminator
4177 .. 4884  =  708 bp
octopine synthase terminator
pVS1 StaA
6452 .. 7081  =  630 bp
209 amino acids  =  22.1 kDa
Product: stability protein from Pseudomonas
plasmid pVS1
pVS1 StaA
6452 .. 7081  =  630 bp
209 amino acids  =  22.1 kDa
Product: stability protein from Pseudomonas
plasmid pVS1
ori
9513 .. 10,101  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
9513 .. 10,101  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
BlpR
373 .. 924  =  552 bp
183 amino acids  =  20.6 kDa
Product: phosphinothricin acetyltransferase
confers resistance to bialophos or phosphinothricin
BlpR
373 .. 924  =  552 bp
183 amino acids  =  20.6 kDa
Product: phosphinothricin acetyltransferase
confers resistance to bialophos or phosphinothricin
MAS promoter
930 .. 1310  =  381 bp
mannopine synthase promoter (Velten et al., 1984)
MAS promoter
930 .. 1310  =  381 bp
mannopine synthase promoter (Velten et al., 1984)
CaMV 35S promoter
2305 .. 2650  =  346 bp
strong constitutive promoter from cauliflower
mosaic virus
CaMV 35S promoter
2305 .. 2650  =  346 bp
strong constitutive promoter from cauliflower
mosaic virus
MAS terminator
111 .. 363  =  253 bp
mannopine synthase terminator
MAS terminator
111 .. 363  =  253 bp
mannopine synthase terminator
pVS1 oriV
8649 .. 8843  =  195 bp
origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)
pVS1 oriV
8649 .. 8843  =  195 bp
origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)
bom
9187 .. 9327  =  141 bp
basis of mobility region from pBR322
bom
9187 .. 9327  =  141 bp
basis of mobility region from pBR322
TMV Ω
2680 .. 2734  =  55 bp
translational enhancer from the tobacco mosaic
virus 5'-leader sequence (Gallie et al., 1988)
TMV Ω
2680 .. 2734  =  55 bp
translational enhancer from the tobacco mosaic
virus 5'-leader sequence (Gallie et al., 1988)
MCS 2
4122 .. 4160  =  39 bp
multiple cloning site
MCS 2
4122 .. 4160  =  39 bp
multiple cloning site
LB T-DNA repeat
1 .. 25  =  25 bp
left border repeat from nopaline C58 T-DNA
LB T-DNA repeat
1 .. 25  =  25 bp
left border repeat from nopaline C58 T-DNA
RB T-DNA repeat
5128 .. 5152  =  25 bp
right border repeat from nopaline C58 T-DNA
RB T-DNA repeat
5128 .. 5152  =  25 bp
right border repeat from nopaline C58 T-DNA
MCS 1
2734 .. 2764  =  31 bp
multiple cloning site 1
MCS 1
2734 .. 2764  =  31 bp
multiple cloning site 1
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