pSC-A-amp kan (linearized)

Linearized vector with modified 3'-U overhangs and bound topoisomerase, for efficient TA cloning of PCR products.

Sequence Author: Agilent Technologies

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lac operator BspQI - SapI (3923) AflIII - PciI (3806) AlwNI (3397) BclI * (3046) Bsu36I (2982) AgeI (2932) BstBI (2858) DraIII (2408) SspI (2017) Acc65I (4204) KpnI (4208) PspOMI (4210) EcoO109I (4211) ApaI (4214) AbsI - PaeR7I - PspXI - XhoI (4219) SalI (4225) AccI (4226) HincII (4227) BspDI - ClaI (4235) HindIII (4240) EcoRV (4248) EcoRI (4259) End (4270) Start (1) EcoRI (7) PstI (26) TspMI - XmaI (28) SmaI (30) BamHI (34) SpeI (40) XbaI (46) NotI (53) SacII (64) Eco53kI (69) SacI (71) RsrII (483) NcoI - StyI (566) SphI (601) MscI (920) KasI (999) NarI (1000) SfoI (1001) PluTI (1003) AhdI (1213) BsaI (1274) BpmI (1283) PvuI (1583) XmnI (1812) pSC-A-amp/kan 4269 bp
BspQI  (3923)
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  (3923)
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.
AflIII  (3806)
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  (3806)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
AlwNI  (3397)
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.
BclI  (3046)
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.
Bsu36I  (2982)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
AgeI  (2932)
1 site
A C C G G T T G G C C A
BstBI  (2858)
1 site
T T C G A A A A G C T T
DraIII  (2408)
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.
SspI  (2017)
1 site
A A T A T T T T A T A A
Acc65I  (4204)
1 site
G G T A C C C C A T G G
KpnI  (4208)
1 site
G G T A C C C C A T G G
PspOMI  (4210)
1 site
G G G C C C C C C G G G
EcoO109I  (4211)
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.
ApaI  (4214)
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  (4219)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (4219)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (4219)
1 site
V C T C G A G B B G A G C T C V
XhoI  (4219)
1 site
C T C G A G G A G C T C
SalI  (4225)
1 site
G T C G A C C A G C T G
AccI  (4226)
1 site
G T M K A C C A K M T G

Efficient cleavage with AccI requires ≥13 bp on each side of the recognition sequence.
Sticky ends from different AccI sites may not be compatible.
HincII  (4227)
1 site
G T Y R A C C A R Y T G
BspDI  (4235)
1 site
A T C G A T T A G C T A
ClaI  (4235)
1 site
A T C G A T T A G C T A
HindIII  (4240)
1 site
A A G C T T T T C G A A
EcoRV  (4248)
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.
EcoRI  (4259)
2 sites
G A A T T C C T T A A G
End  (4270)
0 sites
Start  (1)
0 sites
EcoRI  (7)
2 sites
G A A T T C C T T A A G
PstI  (26)
1 site
C T G C A G G A C G T C
TspMI  (28)
1 site
C C C G G G G G G C C C
XmaI  (28)
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  (30)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
BamHI  (34)
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  (40)
1 site
A C T A G T T G A T C A
XbaI  (46)
1 site
T C T A G A A G A T C T
NotI  (53)
1 site
G C G G C C G C C G C C G G C G
SacII  (64)
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.
Eco53kI  (69)
1 site
G A G C T C C T C G A G
SacI  (71)
1 site
G A G C T C C T C G A G
RsrII  (483)
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.
NcoI  (566)
1 site
C C A T G G G G T A C C
StyI  (566)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
SphI  (601)
1 site
G C A T G C C G T A C G
MscI  (920)
1 site
T G G C C A A C C G G T
KasI  (999)
1 site
G G C G C C C C G C G G
NarI  (1000)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI recognition sequence.
SfoI  (1001)
1 site
G G C G C C C C G C G G
PluTI  (1003)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI recognition sequence.
AhdI  (1213)
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.
BsaI  (1274)
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.
BpmI  (1283)
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.
PvuI  (1583)
1 site
C G A T C G G C T A G C
XmnI  (1812)
1 site
G A A N N N N T T C C T T N N N N A A G
AmpR
1140 .. 2000  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   1140 .. 1931  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1140 .. 2000  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   1932 .. 2000  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1140 .. 2000  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
NeoR/KanR
337 .. 1131  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
337 .. 1131  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)