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Plasmid Files

pGreenII 0049

Agrobacterium binary vector with kanamycin-resistance and luciferase genes.

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pGreenII 0049 Sequence and MappGreenII 0049.dna
Map and Sequence File   
Sequence Author:  pGreen website
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 DraIII (6939) NruI (6748) AsiSI (6407) Bpu10I - BsmBI (6385) PflMI (6145) AlwNI (5618) ApaLI (5516) BglII (5197) PshAI (4925) BmgBI (4862) PfoI * (4618) BsiWI (4572) AanI (4405) BsrGI (4236) BstEII (4118) XcmI (3996) PpuMI (3548) StyI (421) FseI (528) AanI (546) BglII (627) MauBI (702) PciI (816) AflII (896) RsrII (1116) NmeAIII (1413) PflFI - Tth111I (1516) MscI (1553) NheI (1859) BmtI (1863) BlpI (1920) Acc65I (2274) KpnI (2278) PspOMI (2280) ApaI (2284) AbsI - PaeR7I - PspXI - XhoI (2289) SalI (2295) AccI (2296) HindIII (2310) EcoRV (2318) EcoRI (2322) PstI (2332) TspMI - XmaI (2334) SmaI (2336) BamHI (2340) SpeI (2346) XbaI (2352) NotI (2359) AleI - MslI (2370) Eco53kI (2378) SacI (2380) lac operator ScaI (2801) SgrAI (3298) SexAI * (3386) PacI (3408) pGreenII 0049 7065 bp
DraIII  (6939)
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.
NruI  (6748)
1 site
T C G C G A A G C G C T
AsiSI  (6407)
1 site
G C G A T C G C C G C T A G C G
Bpu10I  (6385)
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.
BsmBI  (6385)
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.
PflMI  (6145)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
AlwNI  (5618)
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.
ApaLI  (5516)
1 site
G T G C A C C A C G T G
BglII  (5197)
2 sites
A G A T C T T C T A G A
PshAI  (4925)
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.
BmgBI  (4862)
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.
PfoI  (4618)
1 site
T C C N G G A A G G N C C T
* Blocked by Dcm methylation.
Sticky ends from different PfoI sites may not be compatible.
BsiWI  (4572)
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.
AanI  (4405)
2 sites
T T A T A A A A T A T T
BsrGI  (4236)
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.
BstEII  (4118)
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.
XcmI  (3996)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
PpuMI  (3548)
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.
StyI  (421)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
FseI  (528)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
AanI  (546)
2 sites
T T A T A A A A T A T T
BglII  (627)
2 sites
A G A T C T T C T A G A
MauBI  (702)
1 site
C G C G C G C G G C G C G C G C
PciI  (816)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AflII  (896)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
RsrII  (1116)
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.
NmeAIII  (1413)
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).
PflFI  (1516)
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  (1516)
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.
MscI  (1553)
1 site
T G G C C A A C C G G T
NheI  (1859)
1 site
G C T A G C C G A T C G
BmtI  (1863)
1 site
G C T A G C C G A T C G
BlpI  (1920)
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.
Acc65I  (2274)
1 site
G G T A C C C C A T G G
KpnI  (2278)
1 site
G G T A C C C C A T G G
PspOMI  (2280)
1 site
G G G C C C C C C G G G
ApaI  (2284)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
AbsI  (2289)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (2289)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (2289)
1 site
V C T C G A G B B G A G C T C V
XhoI  (2289)
1 site
C T C G A G G A G C T C
SalI  (2295)
1 site
G T C G A C C A G C T G
AccI  (2296)
1 site
G T M K A C C A K M T G

Efficient cleavage with AccI requires ≥13 bp on each side of the
recognition sequence.
Sticky ends from different AccI sites may not be compatible.
HindIII  (2310)
1 site
A A G C T T T T C G A A
EcoRV  (2318)
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.
EcoRI  (2322)
1 site
G A A T T C C T T A A G
PstI  (2332)
1 site
C T G C A G G A C G T C
TspMI  (2334)
1 site
C C C G G G G G G C C C
XmaI  (2334)
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  (2336)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
BamHI  (2340)
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.
SpeI  (2346)
1 site
A C T A G T T G A T C A
XbaI  (2352)
1 site
T C T A G A A G A T C T
NotI  (2359)
1 site
G C G G C C G C C G C C G G C G
AleI  (2370)
1 site
C A C N N N N G T G G T G N N N N C A C
MslI  (2370)
1 site
C A Y N N N N R T G G T R N N N N Y A C
Eco53kI  (2378)
1 site
G A G C T C C T C G A G
SacI  (2380)
1 site
G A G C T C C T C G A G
ScaI  (2801)
1 site
A G T A C T T C A T G A
SgrAI  (3298)
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.
SexAI  (3386)
1 site
A C C W G G T T G G W C C A
* Blocked by Dcm methylation.
Sticky ends from different SexAI sites may not be compatible.
PacI  (3408)
1 site
T T A A T T A A A A T T A A T T
luciferase
3079 .. 4731  =  1653 bp
550 amino acids  =  60.7 kDa
Product: firefly luciferase
luciferase
3079 .. 4731  =  1653 bp
550 amino acids  =  60.7 kDa
Product: firefly luciferase
KanR
6022 .. 6837  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
KanR
6022 .. 6837  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
NeoR/KanR
970 .. 1764  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
970 .. 1764  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
ori
5263 .. 5851  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
5263 .. 5851  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
pSa ori
63 .. 498  =  436 bp
origin of replication from bacterial plasmid pSa
pSa ori
63 .. 498  =  436 bp
origin of replication from bacterial plasmid pSa
lacZα
2048 .. 2437  =  390 bp
129 amino acids  =  14.2 kDa
Product: LacZα fragment of β-galactosidase
lacZα
2048 .. 2437  =  390 bp
129 amino acids  =  14.2 kDa
Product: LacZα fragment of β-galactosidase
CaMV 35S promoter
4742 .. 5087  =  346 bp
strong constitutive promoter from cauliflower
mosaic virus
CaMV 35S promoter
4742 .. 5087  =  346 bp
strong constitutive promoter from cauliflower
mosaic virus
NOS terminator
679 .. 930  =  252 bp
nopaline synthase terminator and poly(A) signal
NOS terminator
679 .. 930  =  252 bp
nopaline synthase terminator and poly(A) signal
NOS promoter
1798 .. 1977  =  180 bp
nopaline synthase promoter
NOS promoter
1798 .. 1977  =  180 bp
nopaline synthase promoter
CaMV poly(A) signal
2787 .. 2963  =  177 bp
cauliflower mosaic virus polyadenylation signal
CaMV poly(A) signal
2787 .. 2963  =  177 bp
cauliflower mosaic virus polyadenylation signal
lac promoter
2481 .. 2512  =  32 bp
   Segment 3:  -10  
   2481 .. 2487  =  7 bp
promoter for the E. coli lac operon
lac promoter
2481 .. 2512  =  32 bp
   Segment 2:  
   2488 .. 2506  =  19 bp
promoter for the E. coli lac operon
lac promoter
2481 .. 2512  =  32 bp
   Segment 1:  -35  
   2507 .. 2512  =  6 bp
promoter for the E. coli lac operon
lac promoter
2481 .. 2512  =  32 bp
3 segments
promoter for the E. coli lac operon
RB T-DNA repeat
5148 .. 5172  =  25 bp
right border repeat from nopaline C58 T-DNA
RB T-DNA repeat
5148 .. 5172  =  25 bp
right border repeat from nopaline C58 T-DNA
LB T-DNA repeat
633 .. 655  =  23 bp
left border repeat from nopaline C58 T-DNA
(truncated)
LB T-DNA repeat
633 .. 655  =  23 bp
left border repeat from nopaline C58 T-DNA
(truncated)
lac operator
2457 .. 2473  =  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
2457 .. 2473  =  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).
MCS
2274 .. 2381  =  108 bp
pBluescript multiple cloning site
MCS
2274 .. 2381  =  108 bp
pBluescript multiple cloning site
T7 promoter
2247 .. 2265  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
2247 .. 2265  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T3 promoter
2394 .. 2412  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T3 promoter
2394 .. 2412  =  19 bp
promoter for bacteriophage T3 RNA polymerase
M13 fwd
2224 .. 2240  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
2224 .. 2240  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2433 .. 2449  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2433 .. 2449  =  17 bp
common sequencing primer, one of multiple similar
variants
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