pCR-XL-TOPO (linearized)

Linearized vector with 3'-T overhangs and bound topoisomerase, for TOPO® TA cloning of long PCR products. Positive clones disrupt a lethal lacZα-ccdB fusion gene.

Sequence Author: Thermo Fisher (Invitrogen)

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HomePlasmidsTA and GC Cloning VectorspCR-XL-TOPO (linearized)
PciI (3058) DraI (2303) DraIII (2255) FseI (2178) CsiI - SexAI * (2065) SgrAI (2016) SmaI (1992) TspMI - XmaI (1990) AatII (1948) ZraI (1946) BssHII - MauBI (1938) HincII (1912) SfiI (1902) TsoI (1558) RsrII (1545) MluI (3431) HindIII (3459) Acc65I (3465) KpnI (3469) Eco53kI (3473) SacI (3475) BamHI (3477) SpeI (3483) End (3519) Start (1) PstI (16) EcoRV (19) NotI (34) PaeR7I - PspXI - XhoI (40) XbaI (52) EcoO109I - PspOMI (58) ApaI (62) PvuI (213) BsiWI - SnaBI (243) BsrGI - TatI (303) BsaI (468) XcmI (845) BclI * (870) PflFI - Tth111I (1147) BsrDI (1262) pCR®-XL-TOPO® 3518 bp
PciI  (3058)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
DraI  (2303)
1 site		 T T T A A A A A A T T T
DraIII  (2255)
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.
FseI  (2178)
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.
CsiI  (2065)
1 site		 A C C W G G T T G G W C C A
Sticky ends from different CsiI sites may not be compatible.
SexAI  (2065)
1 site		 A C C W G G T T G G W C C A
* Blocked by Dcm methylation.
Sticky ends from different SexAI sites may not be compatible.
SgrAI  (2016)
1 site		 C R C C G G Y G G Y G G C C R C
Efficient cleavage requires at least two copies of the SgrAI recognition sequence.
SmaI  (1992)
1 site		 C C C G G G G G G C C C
SmaI can be used at 37°C for brief incubations.
TspMI  (1990)
1 site		 C C C G G G G G G C C C
XmaI  (1990)
1 site		 C C C G G G G G G C C C
Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
AatII  (1948)
1 site		 G A C G T C C T G C A G
ZraI  (1946)
1 site		 G A C G T C C T G C A G
BssHII  (1938)
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.
MauBI  (1938)
1 site		 C G C G C G C G G C G C G C G C
HincII  (1912)
1 site		 G T Y R A C C A R Y T G
SfiI  (1902)
1 site		 G G C C N N N N N G G C C C C G G N N N N N C C G G
Efficient cleavage requires at least two copies of the SfiI recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
TsoI  (1558)
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).
RsrII  (1545)
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.
MluI  (3431)
1 site		 A C G C G T T G C G C A
HindIII  (3459)
1 site		 A A G C T T T T C G A A
Acc65I  (3465)
1 site		 G G T A C C C C A T G G
KpnI  (3469)
1 site		 G G T A C C C C A T G G
Eco53kI  (3473)
1 site		 G A G C T C C T C G A G
SacI  (3475)
1 site		 G A G C T C C T C G A G
BamHI  (3477)
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  (3483)
1 site		 A C T A G T T G A T C A
End  (3519)
0 sites
Start  (1)
0 sites
PstI  (16)
1 site		 C T G C A G G A C G T C
EcoRV  (19)
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  (34)
1 site		 G C G G C C G C C G C C G G C G
PaeR7I  (40)
1 site		 C T C G A G G A G C T C
PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (40)
1 site		 V C T C G A G B B G A G C T C V
XhoI  (40)
1 site		 C T C G A G G A G C T C
XbaI  (52)
1 site		 T C T A G A A G A T C T
EcoO109I  (58)
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  (58)
1 site		 G G G C C C C C C G G G
ApaI  (62)
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  (213)
1 site		 C G A T C G G C T A G C
BsiWI  (243)
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  (243)
1 site		 T A C G T A A T G C A T
BsrGI  (303)
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  (303)
1 site		 W G T A C W W C A T G W
BsaI  (468)
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.
XcmI  (845)
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  (870)
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  (1147)
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  (1147)
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.
BsrDI  (1262)
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
901 .. 1695  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
901 .. 1695  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
ori
2414 .. 3002  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2414 .. 3002  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
LacZα
2 .. 240  =  239 bp
79 amino acids  =  9.2 kDa
Product: LacZα fragment of β-galactosidase
LacZα
2 .. 240  =  239 bp
79 amino acids  =  9.2 kDa
Product: LacZα fragment of β-galactosidase
ccdB
250 .. 552  =  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
250 .. 552  =  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.
BleoR
1902 .. 2276  =  375 bp
124 amino acids  =  13.8 kDa
Product: antibiotic-binding protein
confers resistance to bleomycin, phleomycin, and Zeocin™
BleoR
1902 .. 2276  =  375 bp
124 amino acids  =  13.8 kDa
Product: antibiotic-binding protein
confers resistance to bleomycin, phleomycin, and Zeocin™
LacZα
3400 .. 3518  =  119 bp
39 amino acids  =  4.1 kDa
Product: LacZα fragment of β-galactosidase
LacZα
3400 .. 3518  =  119 bp
39 amino acids  =  4.1 kDa
Product: LacZα fragment of β-galactosidase
lac promoter
3326 .. 3356  =  31 bp
3 segments
   Segment 1:  -35  
   3326 .. 3331  =  6 bp
promoter for the E. coli lac operon
lac promoter
3326 .. 3356  =  31 bp
3 segments
   Segment 2:  
   3332 .. 3349  =  18 bp
promoter for the E. coli lac operon
lac promoter
3326 .. 3356  =  31 bp
3 segments
   Segment 3:  -10  
   3350 .. 3356  =  7 bp
promoter for the E. coli lac operon
lac promoter
3326 .. 3356  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
3364 .. 3380  =  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
3364 .. 3380  =  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).
T7 promoter
71 .. 89  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
71 .. 89  =  19 bp
promoter for bacteriophage T7 RNA polymerase
M13 fwd
96 .. 112  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
96 .. 112  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
3388 .. 3404  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
3388 .. 3404  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  1 .. 552  =  552 bp
ORF:  183 amino acids  =  21.3 kDa  (no start codon)
ORF:  901 .. 1695  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  2 .. 346  =  345 bp
ORF:  114 amino acids  =  12.6 kDa  (no start codon)
ORF:  1073 .. 1459  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  1902 .. 2276  =  375 bp
ORF:  124 amino acids  =  13.8 kDa
ORF:  1210 .. 1743  =  534 bp
ORF:  177 amino acids  =  19.9 kDa
ORF:  1874 .. 2182  =  309 bp
ORF:  102 amino acids  =  10.9 kDa
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Individual Sequences & Maps

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pTZ57R_T
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pcDNA5 FRT V5-His-TOPO (linearized)
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T-Vector pMD19 (Simple)
pcDNA6.2 N-EmGFP-GW TOPO (linearized)
pGlow-TOPO (linearized)
pSMART GC LK (linearized)
T-Vector pMD20
pCR-XL-TOPO (linearized)
pIB V5-His-TOPO (linearized)

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