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Plasmid Files

pGreenII 0229

Agrobacterium binary vector with kanamycin-and bialophos/phosphinothricin-resistance genes.

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pGreenII 0229 Sequence and MappGreenII 0229.dna
Map and Sequence File   
Sequence Author:  pGreen website
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 DraIII (4322) BsrDI (4298) NruI (4131) TsoI (3949) PasI (3913) EcoNI (3875) AsiSI (3790) Bpu10I (3768) PflMI (3528) AlwNI (3001) ApaLI (2899) PspFI (2893) BseYI (2889) BglII (2580) StuI (2527) BspQI - SapI (2403) lac operator StyI (421) FseI (528) AanI - PsiI (546) BglII (627) LB T-DNA repeat MauBI (702) AflIII - PciI (816) AflII (896) BmgBI (1021) FspAI (1098) AarI - BfuAI - BspMI (1162) PpuMI - SanDI (1275) AfeI (1335) PflFI - Tth111I (1352) BsgI (1455) BsaHI (1484) NheI (1605) BmtI (1609) Bsu36I (1621) BlpI (1666) Acc65I (2018) KpnI (2022) PspOMI (2024) ApaI (2028) AbsI - PaeR7I - PspXI - XhoI (2033) SalI (2039) AccI (2040) HindIII (2054) EcoRV (2062) EcoRI (2066) PstI (2076) TspMI - XmaI (2078) SmaI (2080) BamHI (2084) SpeI (2090) XbaI (2096) NotI (2103) AleI - MslI (2114) SacII (2115) BstXI (2116) Eco53kI (2122) SacI (2124) pGreenII 0229 4448 bp
DraIII  (4322)
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.
BsrDI  (4298)
1 site
G C A A T G N N C G T T A C

Sticky ends from different BsrDI sites may not be compatible.
NruI  (4131)
1 site
T C G C G A A G C G C T
TsoI  (3949)
1 site
T A R C C A ( N ) 9 N N A T Y G G T ( N ) 9

Sticky ends from different TsoI sites may not be compatible.
After cleavage, TsoI can remain bound to DNA and alter its
electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
PasI  (3913)
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.
EcoNI  (3875)
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.
AsiSI  (3790)
1 site
G C G A T C G C C G C T A G C G
Bpu10I  (3768)
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.
PflMI  (3528)
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  (3001)
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  (2899)
1 site
G T G C A C C A C G T G
PspFI  (2893)
1 site
C C C A G C G G G T C G
BseYI  (2889)
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.
BglII  (2580)
2 sites
A G A T C T T C T A G A
StuI  (2527)
1 site
A G G C C T T C C G G A
BspQI  (2403)
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  (2403)
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.
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)
1 site
T T A T A A A A T A T T
PsiI  (546)
1 site
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
AflIII  (816)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
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.
BmgBI  (1021)
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.
FspAI  (1098)
1 site
R T G C G C A Y Y A C G C G T R
AarI  (1162)
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.
BfuAI  (1162)
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  (1162)
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.
PpuMI  (1275)
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.
SanDI  (1275)
1 site
G G G W C C C C C C W G G G

Sticky ends from different SanDI sites may not be compatible.
AfeI  (1335)
1 site
A G C G C T T C G C G A
PflFI  (1352)
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  (1352)
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.
BsgI  (1455)
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).
BsaHI  (1484)
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.
NheI  (1605)
1 site
G C T A G C C G A T C G
BmtI  (1609)
1 site
G C T A G C C G A T C G
Bsu36I  (1621)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
BlpI  (1666)
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  (2018)
1 site
G G T A C C C C A T G G
KpnI  (2022)
1 site
G G T A C C C C A T G G
PspOMI  (2024)
1 site
G G G C C C C C C G G G
ApaI  (2028)
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  (2033)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (2033)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (2033)
1 site
V C T C G A G B B G A G C T C V
XhoI  (2033)
1 site
C T C G A G G A G C T C
SalI  (2039)
1 site
G T C G A C C A G C T G
AccI  (2040)
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  (2054)
1 site
A A G C T T T T C G A A
EcoRV  (2062)
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  (2066)
1 site
G A A T T C C T T A A G
PstI  (2076)
1 site
C T G C A G G A C G T C
TspMI  (2078)
1 site
C C C G G G G G G C C C
XmaI  (2078)
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  (2080)
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  (2084)
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  (2090)
1 site
A C T A G T T G A T C A
XbaI  (2096)
1 site
T C T A G A A G A T C T
NotI  (2103)
1 site
G C G G C C G C C G C C G G C G
AleI  (2114)
1 site
C A C N N N N G T G G T G N N N N C A C
MslI  (2114)
1 site
C A Y N N N N R T G G T R N N N N Y A C
SacII  (2115)
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  (2116)
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.
Eco53kI  (2122)
1 site
G A G C T C C T C G A G
SacI  (2124)
1 site
G A G C T C C T C G A G
KanR
3405 .. 4220  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
KanR
3405 .. 4220  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
ori
2646 .. 3234  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2646 .. 3234  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
BlpR
950 .. 1501  =  552 bp
183 amino acids  =  20.6 kDa
Product: phosphinothricin acetyltransferase
confers resistance to bialophos or phosphinothricin
BlpR
950 .. 1501  =  552 bp
183 amino acids  =  20.6 kDa
Product: phosphinothricin acetyltransferase
confers resistance to bialophos or phosphinothricin
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
1792 .. 2181  =  390 bp
129 amino acids  =  14.2 kDa
Product: LacZα fragment of β-galactosidase
lacZα
1792 .. 2181  =  390 bp
129 amino acids  =  14.2 kDa
Product: LacZα fragment of β-galactosidase
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
1544 .. 1721  =  178 bp
nopaline synthase promoter
NOS promoter
1544 .. 1721  =  178 bp
nopaline synthase promoter
lac promoter
2225 .. 2256  =  32 bp
   Segment 3:  -10  
   2225 .. 2231  =  7 bp
promoter for the E. coli lac operon
lac promoter
2225 .. 2256  =  32 bp
   Segment 2:  
   2232 .. 2250  =  19 bp
promoter for the E. coli lac operon
lac promoter
2225 .. 2256  =  32 bp
   Segment 1:  -35  
   2251 .. 2256  =  6 bp
promoter for the E. coli lac operon
lac promoter
2225 .. 2256  =  32 bp
3 segments
promoter for the E. coli lac operon
RB T-DNA repeat
2531 .. 2555  =  25 bp
right border repeat from nopaline C58 T-DNA
RB T-DNA repeat
2531 .. 2555  =  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
2201 .. 2217  =  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
2201 .. 2217  =  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
2018 .. 2125  =  108 bp
pBluescript multiple cloning site
MCS
2018 .. 2125  =  108 bp
pBluescript multiple cloning site
T7 promoter
1991 .. 2009  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
1991 .. 2009  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T3 promoter
2138 .. 2156  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T3 promoter
2138 .. 2156  =  19 bp
promoter for bacteriophage T3 RNA polymerase
M13 fwd
1968 .. 1984  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
1968 .. 1984  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2177 .. 2193  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2177 .. 2193  =  17 bp
common sequencing primer, one of multiple similar
variants
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