pBluescript II KS(+)

Standard cloning vector (phagemid excised from lambda ZAPII). The f1 (+) orientation allows rescue of sense strand ssDNA. pBluescript II SK(+) has a reversed MCS.
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XmnI (2645) BsaHI (2583) ScaI (2526) TatI (2524) NmeAIII (2194) BpmI (2116) BsaI (2107) AhdI (2046) AlwNI (1569) PsiI (99) BsaAI - DraIII (227) BtgZI (228) NgoMIV (328) NaeI (330) Eco53kI (655) SacI (657) BtgI (661) AleI (663) SacII (664) BstXI (665) EagI - NotI (670) XbaI (677) SpeI (683) BamHI (689) TspMI - XmaI (695) SmaI (697) PstI (705) EcoRI (707) EcoRV (715) HindIII (719) BspDI - ClaI (726) SalI (734) AccI (735) HincII (736) AbsI - PaeR7I - PspXI - XhoI (740) EcoO109I - PspOMI (749) ApaI (753) Acc65I (755) KpnI (759) BspQI - SapI (1037) AflIII - PciI (1153) NspI (1157) pBluescript II KS(+) 2961 bp
XmnI  (2645)
1 site
G A A N N N N T T C C T T N N N N A A G
BsaHI  (2583)
1 site
G R C G Y C C Y G C R G

BsaHI is typically used at 37°C, but is even more active at 60°C.
ScaI  (2526)
1 site
A G T A C T T C A T G A
TatI  (2524)
1 site
W G T A C W W C A T G W
NmeAIII  (2194)
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).
BpmI  (2116)
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.
BsaI  (2107)
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  (2046)
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  (1569)
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.
PsiI  (99)
1 site
T T A T A A A A T A T T
BsaAI  (227)
1 site
Y A C G T R R T G C A Y
DraIII  (227)
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.
BtgZI  (228)
1 site
G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4

Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
NgoMIV  (328)
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  (330)
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.
Eco53kI  (655)
1 site
G A G C T C C T C G A G
SacI  (657)
1 site
G A G C T C C T C G A G
BtgI  (661)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
AleI  (663)
1 site
C A C N N N N G T G G T G N N N N C A C
SacII  (664)
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.
BstXI  (665)
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.
EagI  (670)
1 site
C G G C C G G C C G G C
NotI  (670)
1 site
G C G G C C G C C G C C G G C G
XbaI  (677)
1 site
T C T A G A A G A T C T
SpeI  (683)
1 site
A C T A G T T G A T C A
BamHI  (689)
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.
TspMI  (695)
1 site
C C C G G G G G G C C C
XmaI  (695)
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  (697)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
PstI  (705)
1 site
C T G C A G G A C G T C
EcoRI  (707)
1 site
G A A T T C C T T A A G
EcoRV  (715)
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.
HindIII  (719)
1 site
A A G C T T T T C G A A
BspDI  (726)
1 site
A T C G A T T A G C T A
ClaI  (726)
1 site
A T C G A T T A G C T A
SalI  (734)
1 site
G T C G A C C A G C T G
AccI  (735)
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  (736)
1 site
G T Y R A C C A R Y T G
AbsI  (740)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (740)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (740)
1 site
V C T C G A G B B G A G C T C V
XhoI  (740)
1 site
C T C G A G G A G C T C
EcoO109I  (749)
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  (749)
1 site
G G G C C C C C C G G G
ApaI  (753)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
Acc65I  (755)
1 site
G G T A C C C C A T G G
KpnI  (759)
1 site
G G T A C C C C A T G G
BspQI  (1037)
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  (1037)
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  (1153)
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  (1153)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
NspI  (1157)
1 site
R C A T G Y Y G T A C R
AmpR
1973 .. 2833  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   1973 .. 2764  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1973 .. 2833  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   2765 .. 2833  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1973 .. 2833  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
1214 .. 1802  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
1214 .. 1802  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
lacZα
241 .. 816  =  576 bp
191 amino acids  =  20.8 kDa
Product: LacZα fragment of β-galactosidase
lacZα
241 .. 816  =  576 bp
191 amino acids  =  20.8 kDa
Product: LacZα fragment of β-galactosidase
AmpR promoter
2834 .. 2938  =  105 bp
AmpR promoter
2834 .. 2938  =  105 bp
lac promoter
860 .. 890  =  31 bp
3 segments
   Segment 3:  -10  
   860 .. 866  =  7 bp
promoter for the E. coli lac operon
lac promoter
860 .. 890  =  31 bp
3 segments
   Segment 2:  
   867 .. 884  =  18 bp
promoter for the E. coli lac operon
lac promoter
860 .. 890  =  31 bp
3 segments
   Segment 1:  -35  
   885 .. 890  =  6 bp
promoter for the E. coli lac operon
lac promoter
860 .. 890  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
836 .. 852  =  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
836 .. 852  =  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).
f1 ori
3 .. 458  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
3 .. 458  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
MCS
653 .. 760  =  108 bp
pBluescript multiple cloning site
MCS
653 .. 760  =  108 bp
pBluescript multiple cloning site
T7 promoter
626 .. 644  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
626 .. 644  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T3 promoter
773 .. 791  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T3 promoter
773 .. 791  =  19 bp
promoter for bacteriophage T3 RNA polymerase
M13 fwd
600 .. 616  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
600 .. 616  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
812 .. 828  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
812 .. 828  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  2103 .. 2369  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  241 .. 816  =  576 bp
ORF:  191 amino acids  =  20.8 kDa
ORF:  1973 .. 2833  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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