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Plasmid Files

pCR®2.1-TOPO®

Parental bacterial expression vector for TOPO® TA cloning of PCR products.

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pCR2.1-TOPO Sequence and MappCR2.1-TOPO.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 PciI (3806) BspHI (3086) AhdI (2918) BsaI (2852) BpmI (2849) ScaI (2437) TatI (2435) XmnI (2318) CAP binding site lac operator HindIII (234) Acc65I (240) KpnI (244) Eco53kI (248) SacI (250) BamHI (252) SpeI (258) BstXI (279) EcoRI (283) AflII (292) EcoRI (301) EcoRV (313) BstXI (323) NotI (328) AvaI - BsoBI - PaeR7I - PspXI - XhoI (334) BmeT110I (335) NsiI (345) XbaI (346) EcoO109I - PspOMI (352) ApaI (356) M13 fwd DraIII (780) AanI - AanI - PsiI (905) BstAPI (999) XcmI (1263) BglII (1283) BclI * (1288) MscI (1529) PflFI - Tth111I (1565) BssHII (1844) BtgI - NcoI (1879) RsrII (1963) pCR®2.1-TOPO® 3931 bp
PciI  (3806)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspHI  (3086)
1 site
T C A T G A A G T A C T
AhdI  (2918)
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  (2852)
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  (2849)
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.
ScaI  (2437)
1 site
A G T A C T T C A T G A
TatI  (2435)
1 site
W G T A C W W C A T G W
XmnI  (2318)
1 site
G A A N N N N T T C C T T N N N N A A G
HindIII  (234)
1 site
A A G C T T T T C G A A
Acc65I  (240)
1 site
G G T A C C C C A T G G
KpnI  (244)
1 site
G G T A C C C C A T G G
Eco53kI  (248)
1 site
G A G C T C C T C G A G
SacI  (250)
1 site
G A G C T C C T C G A G
BamHI  (252)
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  (258)
1 site
A C T A G T T G A T C A
BstXI  (279)
2 sites
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.
EcoRI  (283)
2 sites
G A A T T C C T T A A G
AflII  (292)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
EcoRI  (301)
2 sites
G A A T T C C T T A A G
EcoRV  (313)
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.
BstXI  (323)
2 sites
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.
NotI  (328)
1 site
G C G G C C G C C G C C G G C G
AvaI  (334)
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  (334)
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.
PaeR7I  (334)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (334)
1 site
V C T C G A G B B G A G C T C V
XhoI  (334)
1 site
C T C G A G G A G C T C
BmeT110I  (335)
1 site
C Y C G R G G R G C Y C
NsiI  (345)
1 site
A T G C A T T A C G T A
XbaI  (346)
1 site
T C T A G A A G A T C T
EcoO109I  (352)
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.
PspOMI  (352)
1 site
G G G C C C C C C G G G
ApaI  (356)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
DraIII  (780)
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.
AanI  (905)
1 site
T T A T A A A A T A T T
AanI  (905)
1 site
T T A T A A A A T A T T
PsiI  (905)
1 site
T T A T A A A A T A T T
BstAPI  (999)
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.
XcmI  (1263)
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.
BglII  (1283)
1 site
A G A T C T T C T A G A
BclI  (1288)
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.
MscI  (1529)
1 site
T G G C C A A C C G G T
PflFI  (1565)
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  (1565)
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.
BssHII  (1844)
1 site
G C G C G C C G C G C G

BssHII is typically used at 50°C, but is 75% active at 37°C.
BtgI  (1879)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
NcoI  (1879)
1 site
C C A T G G G G T A C C
RsrII  (1963)
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.
AmpR
2131 .. 2991  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   2131 .. 2199  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2131 .. 2991  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   2200 .. 2991  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2131 .. 2991  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
NeoR/KanR
1319 .. 2113  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
NeoR/KanR
1319 .. 2113  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin®)
ori
3162 .. 3750  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3162 .. 3750  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
547 .. 975  =  429 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
547 .. 975  =  429 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
lac promoter
143 .. 173  =  31 bp
   Segment 1:  -35  
   143 .. 148  =  6 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
   Segment 2:  
   149 .. 166  =  18 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
   Segment 3:  -10  
   167 .. 173  =  7 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
3 segments
promoter for the E. coli lac operon
CAP binding site
107 .. 128  =  22 bp
CAP binding activates transcription in the presence
of cAMP.
CAP binding site
107 .. 128  =  22 bp
CAP binding activates transcription in the presence
of cAMP.
T7 promoter
365 .. 383  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
365 .. 383  =  19 bp
promoter for bacteriophage T7 RNA polymerase
lac operator
181 .. 197  =  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
181 .. 197  =  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).
M13 rev
205 .. 221  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
205 .. 221  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
390 .. 406  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
390 .. 406  =  17 bp
common sequencing primer, one of multiple similar
variants
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