pCMV SPORT6.1

Gateway® library vector for cloning and transient mammalian cell expression of cDNAs. See pCMV SPORT6 for an older version of this vector.

Sequence Author: Thermo Fisher (Invitrogen)

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BspDI * - ClaI * (123) NaeI (4048) NgoMIV (4046) DraIII (3945) XmnI (3401) ScaI (3282) NmeAIII (2950) BsaI (2863) AhdI (2802) AlwNI (2325) PciI (1909) SV40 poly(A) signal HincII - HpaI (259) MfeI (268) BsaBI * (360) BclI * (361) NheI (637) BmtI (641) M13 fwd T7 promoter MluI (736) HindIII (742) PspOMI (748) ApaI (752) AbsI - PaeR7I - PspXI - XhoI (754) XbaI (766) EagI - NotI (773) TspMI - XmaI (784) SmaI (786) EcoRI (789) RsrII (796) AgeI (798) Acc65I (801) KpnI (805) attB1 SP6 promoter StuI (853) M13 rev SacII (900) AvrII (903) BbsI (932) BsmBI - Esp3I (977) Eco53kI (1004) SacI (1006) CMV promoter NcoI (1206) SnaBI (1230) NsiI (1517) BstAPI (1733) BspQI - SapI (1793) pCMV SPORT6.1 4177 bp
BspDI  (123)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (123)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
NaeI  (4048)
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.
NgoMIV  (4046)
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.
DraIII  (3945)
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.
XmnI  (3401)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (3282)
1 site
A G T A C T T C A T G A
NmeAIII  (2950)
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).
BsaI  (2863)
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.
AhdI  (2802)
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.
AlwNI  (2325)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
PciI  (1909)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
HincII  (259)
1 site
G T Y R A C C A R Y T G
HpaI  (259)
1 site
G T T A A C C A A T T G
MfeI  (268)
1 site
C A A T T G G T T A A C
BsaBI  (360)
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.
BclI  (361)
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.
NheI  (637)
1 site
G C T A G C C G A T C G
BmtI  (641)
1 site
G C T A G C C G A T C G
MluI  (736)
1 site
A C G C G T T G C G C A
HindIII  (742)
1 site
A A G C T T T T C G A A
PspOMI  (748)
1 site
G G G C C C C C C G G G
ApaI  (752)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
AbsI  (754)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (754)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (754)
1 site
V C T C G A G B B G A G C T C V
XhoI  (754)
1 site
C T C G A G G A G C T C
XbaI  (766)
1 site
T C T A G A A G A T C T
EagI  (773)
1 site
C G G C C G G C C G G C
NotI  (773)
1 site
G C G G C C G C C G C C G G C G
TspMI  (784)
1 site
C C C G G G G G G C C C
XmaI  (784)
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.
SmaI  (786)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
EcoRI  (789)
1 site
G A A T T C C T T A A G
RsrII  (796)
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.
AgeI  (798)
1 site
A C C G G T T G G C C A
Acc65I  (801)
1 site
G G T A C C C C A T G G
KpnI  (805)
1 site
G G T A C C C C A T G G
StuI  (853)
1 site
A G G C C T T C C G G A
SacII  (900)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII recognition sequence.
AvrII  (903)
1 site
C C T A G G G G A T C C
BbsI  (932)
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.
BsmBI  (977)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different BsmBI sites may not be compatible.
BsmBI-v2 is an improved version of BsmBI.
Esp3I  (977)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different Esp3I sites may not be compatible.
Eco53kI  (1004)
1 site
G A G C T C C T C G A G
SacI  (1006)
1 site
G A G C T C C T C G A G
NcoI  (1206)
1 site
C C A T G G G G T A C C
SnaBI  (1230)
1 site
T A C G T A A T G C A T
NsiI  (1517)
1 site
A T G C A T T A C G T A
BstAPI  (1733)
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.
BspQI  (1793)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (1793)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be mixed before removing an aliquot.
AmpR
2729 .. 3589  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   2729 .. 3520  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2729 .. 3589  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   3521 .. 3589  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2729 .. 3589  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
1970 .. 2558  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
1970 .. 2558  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
3721 .. 4176  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
3721 .. 4176  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
CMV enhancer
1207 .. 1510  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
1207 .. 1510  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
1003 .. 1206  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
1003 .. 1206  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
SV40 poly(A) signal
125 .. 259  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
125 .. 259  =  135 bp
SV40 polyadenylation signal
AmpR promoter
3590 .. 3694  =  105 bp
AmpR promoter
3590 .. 3694  =  105 bp
attB2
711 .. 735  =  25 bp
recombination site for the Gateway® BP reaction
attB2
711 .. 735  =  25 bp
recombination site for the Gateway® BP reaction
attB1
807 .. 831  =  25 bp
recombination site for the Gateway® BP reaction
attB1
807 .. 831  =  25 bp
recombination site for the Gateway® BP reaction
T7 promoter
691 .. 709  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
691 .. 709  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SP6 promoter
832 .. 850  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
SP6 promoter
832 .. 850  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
M13 fwd
664 .. 680  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
664 .. 680  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
861 .. 877  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
861 .. 877  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  858 .. 1124  =  267 bp
ORF:  88 amino acids  =  9.5 kDa
ORF:  2859 .. 3125  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  1313 .. 1570  =  258 bp
ORF:  85 amino acids  =  9.6 kDa
ORF:  2729 .. 3589  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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