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Plasmid Files

pENTR™4

Gateway® Dual Selection Vector for creating an entry vector by restriction cloning. Same MCS as pENTR™1A except with an NcoI site.

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pENTR4.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Thermo Fisher (Invitrogen)
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NspI (3755) AflIII - PciI (3751) DrdI (3649) BssS α I (3578) BciVI (3553) HaeII (3511) AcuI (3209) AseI (2832) AsiSI - PvuI (2633) EcoNI (2545) NruI (2290) BspHI (2157) EcoRV (1979) PaeR7I - XhoI (1972) BfuAI - BspMI (1938) NheI (44) BmtI (48) ZraI (64) AatII (66) BbsI (134) BmrI (141) BsrBI (199) AflII (343) XmnI (471) Acc65I (491) KpnI (495) BspEI (820) BpmI (946) PasI (1056) ScaI (1241) BssHII (1317) BstZ17I (1358) BbvCI (1548) TspMI - XmaI (1692) SmaI - SrfI (1694) BsaBI * (1707) BmgBI (1728) BstXI (1811) BsaI (1832) pENTR™4 3757 bp
NspI  (3755)
1 site
R C A T G Y Y G T A C R
AflIII  (3751)
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  (3751)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
DrdI  (3649)
1 site
G A C N N N N N N G T C C T G N N N N N N C A G

Sticky ends from different DrdI sites may not be compatible.
BssSαI  (3578)
1 site
C A C G A G G T G C T C
BciVI  (3553)
1 site
G T A T C C ( N ) 5 N C A T A G G ( N ) 5

The 1-base overhangs produced by BciVI may be hard to ligate.
Sticky ends from different BciVI sites may not be compatible.
HaeII  (3511)
1 site
R G C G C Y Y C G C G R
AcuI  (3209)
1 site
C T G A A G ( N ) 14 N N G A C T T C ( N ) 14

Efficient cleavage requires at least two copies of the AcuI recognition sequence.
Sticky ends from different AcuI sites may not be compatible.
After cleavage, AcuI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
AseI  (2832)
1 site
A T T A A T T A A T T A
AsiSI  (2633)
1 site
G C G A T C G C C G C T A G C G
PvuI  (2633)
1 site
C G A T C G G C T A G C
EcoNI  (2545)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
NruI  (2290)
1 site
T C G C G A A G C G C T
BspHI  (2157)
1 site
T C A T G A A G T A C T
EcoRV  (1979)
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.
PaeR7I  (1972)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (1972)
1 site
C T C G A G G A G C T C
BfuAI  (1938)
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  (1938)
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.
NheI  (44)
1 site
G C T A G C C G A T C G
BmtI  (48)
1 site
G C T A G C C G A T C G
ZraI  (64)
1 site
G A C G T C C T G C A G
AatII  (66)
1 site
G A C G T C C T G C A G
BbsI  (134)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
BmrI  (141)
1 site
A C T G G G ( N ) 4 N T G A C C C ( N ) 4

The 1-base overhangs produced by BmrI may be hard to ligate.
Sticky ends from different BmrI sites may not be compatible.
Unlike most restriction enzymes, BmrI can cleave DNA in the absence of magnesium.
BsrBI  (199)
1 site
C C G C T C G G C G A G

This recognition sequence is asymmetric, so ligating blunt ends generated by BsrBI will not always regenerate a BsrBI site.
BsrBI is typically used at 37°C, but can be used at temperatures up to 50°C.
AflII  (343)
1 site
C T T A A G G A A T T C
XmnI  (471)
1 site
G A A N N N N T T C C T T N N N N A A G
Acc65I  (491)
1 site
G G T A C C C C A T G G
KpnI  (495)
1 site
G G T A C C C C A T G G
BspEI  (820)
1 site
T C C G G A A G G C C T
BpmI  (946)
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.
PasI  (1056)
1 site
C C C W G G G G G G W C C C

Sticky ends from different PasI sites may not be compatible.
ScaI  (1241)
1 site
A G T A C T T C A T G A
BssHII  (1317)
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.
BstZ17I  (1358)
1 site
G T A T A C C A T A T G
BbvCI  (1548)
1 site
C C T C A G C G G A G T C G
TspMI  (1692)
1 site
C C C G G G G G G C C C
XmaI  (1692)
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.
SmaI  (1694)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
SrfI  (1694)
1 site
G C C C G G G C C G G G C C C G
BsaBI  (1707)
1 site
G A T N N N N A T C C T A N N N N T A G
* Blocked by Dam methylation.
BmgBI  (1728)
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  (1811)
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.
BsaI  (1832)
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.
KanR
2208 .. 3017  =  810 bp
269 amino acids  =  30.7 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418 (Geneticin®) in eukaryotes
KanR
2208 .. 3017  =  810 bp
269 amino acids  =  30.7 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418 (Geneticin®) in eukaryotes
CmR
611 .. 1291  =  681 bp
226 amino acids  =  26.6 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
611 .. 1291  =  681 bp
226 amino acids  =  26.6 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
3107 .. 3695  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3107 .. 3695  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ccdB
1611 .. 1916  =  306 bp
101 amino acids  =  11.7 kDa
Product: CcdB, a bacterial toxin that poisons DNA gyrase
Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains.
ccdB
1611 .. 1916  =  306 bp
101 amino acids  =  11.7 kDa
Product: CcdB, a bacterial toxin that poisons DNA gyrase
Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains.
attL1
358 .. 457  =  100 bp
recombination site for the Gateway® LR reaction
attL1
358 .. 457  =  100 bp
recombination site for the Gateway® LR reaction
attL2
1986 .. 2085  =  100 bp
recombination site for the Gateway® LR reaction
attL2
1986 .. 2085  =  100 bp
recombination site for the Gateway® LR reaction
rrnB T1 terminator
103 .. 189  =  87 bp
transcription terminator T1 from the E. coli rrnB gene
rrnB T1 terminator
103 .. 189  =  87 bp
transcription terminator T1 from the E. coli rrnB gene
MCS 1
461 .. 510  =  50 bp
multiple cloning site, part 1
MCS 1
461 .. 510  =  50 bp
multiple cloning site, part 1
MCS 2
1951 .. 1982  =  32 bp
multiple cloning site, part 2
MCS 2
1951 .. 1982  =  32 bp
multiple cloning site, part 2
lac UV5 promoter
527 .. 557  =  31 bp
   Segment 1:  -35  
   527 .. 532  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
527 .. 557  =  31 bp
   Segment 2:  
   533 .. 550  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
527 .. 557  =  31 bp
   Segment 3:  -10  
   551 .. 557  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
527 .. 557  =  31 bp
3 segments
E. coli lac promoter with an "up" mutation
rrnB T2 terminator
281 .. 308  =  28 bp
transcription terminator T2 from the E. coli rrnB gene
rrnB T2 terminator
281 .. 308  =  28 bp
transcription terminator T2 from the E. coli rrnB gene
ORF:  611 .. 1291  =  681 bp
ORF:  226 amino acids  =  26.6 kDa
ORF:  1611 .. 1916  =  306 bp
ORF:  101 amino acids  =  11.7 kDa
ORF:  2208 .. 3017  =  810 bp
ORF:  269 amino acids  =  30.7 kDa
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