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Plasmid Files

pZErO®-2

Vector for high-efficiency cloning of DNA inserts with sticky or blunt ends. A toxic lacZα-ccdB gene selects against clones lacking an insert.

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pZErO-2.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Thermo Fisher (Invitrogen)
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AflIII - PciI (3172) BspHI (2452) XcmI (2172) BglII (2147) BclI * (2142) PflFI - Tth111I (1868) NmeAIII (1765) HindIII (276) Acc65I (282) KpnI (286) Eco53kI (290) SacI (292) BamHI (294) SpeI (300) EcoRI (325) PstI (334) EcoRV (337) NotI (352) AvaI - BsoBI - PaeR7I - PspXI - XhoI (358) BmeT110I (359) XbaI (370) EcoO109I - PspOMI (376) ApaI (380) PvuI (531) BsiWI - SnaBI (561) BsrGI - TatI (621) BmgBI (682) BsaI (786) StuI (878) BglI (884) BssHII (919) DraIII (1127) PsiI (1252) RsrII (1468) BtgI - NcoI (1551) BsrDI (1756) pZErO®-2 3297 bp
AflIII  (3172)
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  (3172)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspHI  (2452)
1 site
T C A T G A A G T A C T
XcmI  (2172)
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  (2147)
1 site
A G A T C T T C T A G A
BclI  (2142)
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.
PflFI  (1868)
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  (1868)
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.
NmeAIII  (1765)
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).
HindIII  (276)
1 site
A A G C T T T T C G A A
Acc65I  (282)
1 site
G G T A C C C C A T G G
KpnI  (286)
1 site
G G T A C C C C A T G G
Eco53kI  (290)
1 site
G A G C T C C T C G A G
SacI  (292)
1 site
G A G C T C C T C G A G
BamHI  (294)
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  (300)
1 site
A C T A G T T G A T C A
EcoRI  (325)
1 site
G A A T T C C T T A A G
PstI  (334)
1 site
C T G C A G G A C G T C
EcoRV  (337)
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.
NotI  (352)
1 site
G C G G C C G C C G C C G G C G
AvaI  (358)
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  (358)
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  (358)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (358)
1 site
V C T C G A G B B G A G C T C V
XhoI  (358)
1 site
C T C G A G G A G C T C
BmeT110I  (359)
1 site
C Y C G R G G R G C Y C
XbaI  (370)
1 site
T C T A G A A G A T C T
EcoO109I  (376)
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  (376)
1 site
G G G C C C C C C G G G
ApaI  (380)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PvuI  (531)
1 site
C G A T C G G C T A G C
BsiWI  (561)
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.
SnaBI  (561)
1 site
T A C G T A A T G C A T
BsrGI  (621)
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.
TatI  (621)
1 site
W G T A C W W C A T G W
BmgBI  (682)
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.
BsaI  (786)
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.
StuI  (878)
1 site
A G G C C T T C C G G A
BglI  (884)
1 site
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
BssHII  (919)
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.
DraIII  (1127)
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.
PsiI  (1252)
1 site
T T A T A A A A T A T T
RsrII  (1468)
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.
BtgI  (1551)
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  (1551)
1 site
C C A T G G G G T A C C
BsrDI  (1756)
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.
NeoR/KanR
1322 .. 2116  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
1322 .. 2116  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
ori
2528 .. 3116  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2528 .. 3116  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
M13 ori
895 .. 1274  =  380 bp
M13 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
M13 ori
895 .. 1274  =  380 bp
M13 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
lacZα
217 .. 558  =  342 bp
114 amino acids  =  12.9 kDa
Product: LacZα fragment of β-galactosidase
lacZα
217 .. 558  =  342 bp
114 amino acids  =  12.9 kDa
Product: LacZα fragment of β-galactosidase
ccdB
568 .. 870  =  303 bp
100 amino acids  =  11.6 kDa
Product: CcdB, a bacterial toxin that poisons DNA gyrase
Plasmids containing the ccdB gene cannot be propagaged in standard E. coli strains.
ccdB
568 .. 870  =  303 bp
100 amino acids  =  11.6 kDa
Product: CcdB, a bacterial toxin that poisons DNA gyrase
Plasmids containing the ccdB gene cannot be propagaged in standard E. coli strains.
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
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).
MCS
269 .. 381  =  113 bp
multiple cloning site
MCS
269 .. 381  =  113 bp
multiple cloning site
SP6 promoter
239 .. 257  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
SP6 promoter
239 .. 257  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
T7 promoter
389 .. 407  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
389 .. 407  =  19 bp
promoter for bacteriophage T7 RNA polymerase
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
414 .. 430  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
414 .. 430  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  217 .. 870  =  654 bp
ORF:  217 amino acids  =  24.9 kDa
ORF:  365 .. 664  =  300 bp
ORF:  99 amino acids  =  11.1 kDa
ORF:  1553 .. 1807  =  255 bp
ORF:  84 amino acids  =  9.6 kDa
ORF:  1558 .. 1944  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  1322 .. 2116  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
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