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Plasmid Files

T-Vector pMD20

Linearized vector with a pUC19-derived MCS and with a single 3'-T at each end, for TA cloning of PCR products and blue-white screening.

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T-Vector pMD20 Sequence and MapT-Vector pMD20.dna
Map and Sequence File   
Sequence Author:  TaKaRa
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 HindIII (2687) BspQI - SapI (2418) AflIII - PciI (2301) PspFI (2001) BseYI (1997) AlwNI (1892) AhdI (1413) BsaI (1347) BpmI (1344) BsrFI (1328) NmeAIII (1266) BfuAI - BspMI (2692) SphI (2697) PstI - SbfI (2703) SalI (2705) AccI (2706) HincII (2707) XbaI (2711) SpeI (2723) NdeI (2730) End (2738) Start (1) BamHI (5) AvaI - BsoBI - TspMI - XmaI (10) BmeT110I (11) SmaI (12) Acc65I (14) KpnI (18) Eco53kI (22) BanII - SacI (24) ApoI - EcoRI (26) BstAPI (243) PfoI (374) EcoO109I (433) ZraI (492) AatII (494) SspI (608) XmnI (813) ScaI (932) TsoI (1015) T-Vector pMD20 2737 bp
HindIII  (2687)
1 site
A A G C T T T T C G A A
BspQI  (2418)
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  (2418)
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.
AflIII  (2301)
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  (2301)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PspFI  (2001)
1 site
C C C A G C G G G T C G
BseYI  (1997)
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  (1892)
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  (1413)
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.
BsaI  (1347)
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.
BpmI  (1344)
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.
BsrFI  (1328)
1 site
R C C G G Y Y G G C C R

Efficient cleavage requires at least two copies of the BsrFI
recognition sequence.
After cleavage, BsrFI can remain bound to DNA and alter its
electrophoretic mobility.
NmeAIII  (1266)
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).
BfuAI  (2692)
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  (2692)
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.
SphI  (2697)
1 site
G C A T G C C G T A C G
PstI  (2703)
1 site
C T G C A G G A C G T C
SbfI  (2703)
1 site
C C T G C A G G G G A C G T C C
SalI  (2705)
1 site
G T C G A C C A G C T G
AccI  (2706)
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.
HincII  (2707)
1 site
G T Y R A C C A R Y T G
XbaI  (2711)
1 site
T C T A G A A G A T C T
SpeI  (2723)
1 site
A C T A G T T G A T C A
NdeI  (2730)
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.
End  (2738)
0 sites
Start  (1)
0 sites
BamHI  (5)
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.
AvaI  (10)
1 site
C Y C G R G G R G C Y C

Sticky ends from different AvaI sites may not be compatible.
BsoBI  (10)
1 site
C Y C G R G G R G C Y C

Sticky ends from different BsoBI sites may not be compatible.
BsoBI is typically used at 37°C, but can be used at temperatures
up to 65°C.
TspMI  (10)
1 site
C C C G G G G G G C C C
XmaI  (10)
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.
BmeT110I  (11)
1 site
C Y C G R G G R G C Y C
SmaI  (12)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
Acc65I  (14)
1 site
G G T A C C C C A T G G
KpnI  (18)
1 site
G G T A C C C C A T G G
Eco53kI  (22)
1 site
G A G C T C C T C G A G
BanII  (24)
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  (24)
1 site
G A G C T C C T C G A G
ApoI  (26)
1 site
R A A T T Y Y T T A A R

ApoI is typically used at 50°C, but is 50% active at 37°C.
EcoRI  (26)
1 site
G A A T T C C T T A A G
BstAPI  (243)
1 site
G C A N N N N N T G C C G T N N N N N A C G

Sticky ends from different BstAPI sites may not be compatible.
PfoI  (374)
1 site
T C C N G G A A G G N C C T

Sticky ends from different PfoI sites may not be compatible.
EcoO109I  (433)
1 site
R G G N C C Y Y C C N G G R

Sticky ends from different EcoO109I sites may not be compatible.
ZraI  (492)
1 site
G A C G T C C T G C A G
AatII  (494)
1 site
G A C G T C C T G C A G
SspI  (608)
1 site
A A T A T T T T A T A A
XmnI  (813)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (932)
1 site
A G T A C T T C A T G A
TsoI  (1015)
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).
AmpR
626 .. 1486  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   626 .. 694  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
626 .. 1486  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   695 .. 1486  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
626 .. 1486  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
1657 .. 2245  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1657 .. 2245  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
lacZα
2 .. 194  =  193 bp
63 amino acids  =  7.2 kDa
Product: LacZα fragment of β-galactosidase
lacZα
2 .. 194  =  193 bp
63 amino acids  =  7.2 kDa
Product: LacZα fragment of β-galactosidase
AmpR promoter
521 .. 625  =  105 bp
AmpR promoter
521 .. 625  =  105 bp
lacZα
2643 .. 2736  =  94 bp
31 amino acids  =  3.5 kDa
Product: LacZα fragment of β-galactosidase
lacZα
2643 .. 2736  =  94 bp
31 amino acids  =  3.5 kDa
Product: LacZα fragment of β-galactosidase
lac promoter
2569 .. 2599  =  31 bp
   Segment 1:  -35  
   2569 .. 2574  =  6 bp
promoter for the E. coli lac operon
lac promoter
2569 .. 2599  =  31 bp
   Segment 2:  
   2575 .. 2592  =  18 bp
promoter for the E. coli lac operon
lac promoter
2569 .. 2599  =  31 bp
   Segment 3:  -10  
   2593 .. 2599  =  7 bp
promoter for the E. coli lac operon
lac promoter
2569 .. 2599  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
2607 .. 2623  =  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
2607 .. 2623  =  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
2687 .. 2736  =  50 bp
multiple cloning site
MCS
2687 .. 2736  =  50 bp
multiple cloning site
MCS
2 .. 31  =  30 bp
multiple cloning site
MCS
2 .. 31  =  30 bp
multiple cloning site
SP6 promoter
2665 .. 2682  =  18 bp
promoter for bacteriophage SP6 RNA polymerase
SP6 promoter
2665 .. 2682  =  18 bp
promoter for bacteriophage SP6 RNA polymerase
M13 fwd
32 .. 48  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
32 .. 48  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2631 .. 2647  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
2631 .. 2647  =  17 bp
common sequencing primer, one of multiple similar
variants
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