pCR4-TOPO

Parental sequencing vector for TOPO® TA cloning of PCR products.

Sequence Author: Thermo Fisher (Invitrogen)

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HomePlasmidsI.M.A.G.E. Consortium PlasmidspCR4-TOPO
AflIII - PciI (3831) SspI (3080) XmnI (2875) ScaI (2756) BglI (2396) BpmI (2346) AhdI (2276) RsrII (1803) lac operator SpeI (262) PstI - SbfI (273) PmeI (278) AflII (292) NotI (308) BsiWI - SnaBI (501) BsrGI (561) BmgBI (622) BstXI (705) XcmI (1103) BclI * (1128) PflFI - Tth111I (1405) BanII (1652) SphI (1692) BtgI - NcoI (1719) NgoMIV (1787) NaeI (1789) pCR®4-TOPO® 3956 bp
AflIII  (3831)
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  (3831)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
SspI  (3080)
1 site		 A A T A T T T T A T A A
XmnI  (2875)
1 site		 G A A N N N N T T C C T T N N N N A A G
ScaI  (2756)
1 site		 A G T A C T T C A T G A
BglI  (2396)
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.
BpmI  (2346)
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.
AhdI  (2276)
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.
RsrII  (1803)
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.
SpeI  (262)
1 site		 A C T A G T T G A T C A
PstI  (273)
1 site		 C T G C A G G A C G T C
SbfI  (273)
1 site		 C C T G C A G G G G A C G T C C
PmeI  (278)
1 site		 G T T T A A A C C A A A T T T G
AflII  (292)
1 site		 C T T A A G G A A T T C
NotI  (308)
1 site		 G C G G C C G C C G C C G G C G
BsiWI  (501)
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  (501)
1 site		 T A C G T A A T G C A T
BsrGI  (561)
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.
BmgBI  (622)
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.
BstXI  (705)
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.
XcmI  (1103)
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.
BclI  (1128)
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  (1405)
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  (1405)
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.
BanII  (1652)
1 site		 G R G C Y C C Y C G R G
Sticky ends from different BanII sites may not be compatible.
SphI  (1692)
1 site		 G C A T G C C G T A C G
BtgI  (1719)
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  (1719)
1 site		 C C A T G G G G T A C C
NgoMIV  (1787)
1 site		 G C C G G C C G G C C G
Efficient cleavage requires at least two copies of the NgoMIV recognition sequence.
NaeI  (1789)
1 site		 G C C G G C C G G C C G
Efficient cleavage requires at least two copies of the NaeI recognition sequence.
AmpR
2203 .. 3063  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   2203 .. 2994  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2203 .. 3063  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   2995 .. 3063  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2203 .. 3063  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
NeoR/KanR
1159 .. 1953  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
1159 .. 1953  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
lacZα
217 .. 498  =  282 bp
94 amino acids  =  10.5 kDa
Product: LacZα fragment of β-galactosidase
lacZα
217 .. 498  =  282 bp
94 amino acids  =  10.5 kDa
Product: LacZα fragment of β-galactosidase
ccdB
508 .. 810  =  303 bp
100 amino acids  =  11.6 kDa
Product: CcdB, a bacterial toxin that poisons DNA gyrase
Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains.
ccdB
508 .. 810  =  303 bp
100 amino acids  =  11.6 kDa
Product: CcdB, a bacterial toxin that poisons DNA gyrase
Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains.
ori
3187 .. 3775  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3187 .. 3775  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
lac promoter
143 .. 173  =  31 bp
3 segments
   Segment 1:  -35  
   143 .. 148  =  6 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
3 segments
   Segment 2:  
   149 .. 166  =  18 bp
promoter for the E. coli lac operon
lac promoter
143 .. 173  =  31 bp
3 segments
   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.
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).
T3 promoter
242 .. 260  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T3 promoter
242 .. 260  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T7 promoter
329 .. 347  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
329 .. 347  =  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
354 .. 370  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
354 .. 370  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  217 .. 810  =  594 bp
ORF:  197 amino acids  =  22.5 kDa
ORF:  1159 .. 1953  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  1331 .. 1717  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  2333 .. 2599  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  1468 .. 2001  =  534 bp
ORF:  177 amino acids  =  19.9 kDa
ORF:  2203 .. 3063  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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Individual Sequences & Maps

Lafmid BA
pcDNAII
pCR2.1-TOPO (linearized)
pExpress-1
pAD-GAL4
pcDNA3.1(+)
pCR3.1
pME18S-FL3
pAMP1
pcDNA3.1(-)
pCR3.1 (linearized)
pOTB7
pAMP10
pcDNA3.1 Zeo(+)
pCR4-TOPO
pOTB7-3
pBK-CMV
pCMV SPORT
pCR4-TOPO (linearized)
pOTB7a
pBluescribe (modified)
pCMV SPORT2
pCS105
pPCR-Script Amp SK(+)
pBluescript II SK(+)
pCMV SPORT4
pCS107
pRKW2
pBluescript KS(+)
pCMV SPORT6
pCS108
pSPORT 1
pBluescript SK(+)
pCMV SPORT6.1
pCS2+
pSPORT 2
pBluescript SK(-)
pCMV SPORT6ccdB
pCS22+
pT7T3D-PacI
pBluescript-FL
pCR-Blunt II-TOPO
pCS2G
pUC19
pBluescript II KS(+)
pCR-Blunt II-TOPO (linearized)
pDNR-Dual
pYX-Asc
pBluescriptR
pCR-XL-TOPO
pDNR-LIB
pZErO-2
pcDNA3
pCR-XL-TOPO (linearized)
pDONR201
pZL1
pcDNAI
pCR2.1-TOPO
pDONR223

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