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Plasmid Files

pSpark® I (linearized)

Linearized high copy vector with an advanced MCS for very efficient cloning of blunt-ended PCR products.

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pSpark I (linearized) Sequence and MappSpark I (linearized).dna
Map and Sequence File   
Sequence Author:  Canvax Biotech
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 T7 promoter NaeI (2653) NgoMIV (2651) BtgZI (2551) BsaAI - DraIII (2550) PsiI (2422) XmnI (1955) BsaHI (1893) ScaI (1836) TatI (1834) NmeAIII (1504) BpmI (1426) BsaI (1417) PspOMI (2971) EcoO109I (2972) ApaI (2975) BtgI - NcoI - StyI (2977) PmeI (2986) EcoRI (2991) BspEI * (2997) StuI (3007) < EcoRV > (3013) < EcoRV > (0) BamHI (5) SpeI (11) BfuAI - BspMI - EagI - NotI (18) PstI - SbfI (29) SalI (31) AccI (32) HincII (33) NdeI (41) Eco53kI (51) AvaI - BsoBI - PaeR7I - PspXI - XhoI (52) BmeT110I - SacI (53) MluI (60) NsiI (73) lac operator BspQI - SapI (347) PciI (463) NspI (467) BseYI (767) PspFI (771) AlwNI (879) AhdI (1356) pSpark® I 3013 bp
NaeI  (2653)
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  (2651)
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.
BtgZI  (2551)
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.
BsaAI  (2550)
1 site
Y A C G T R R T G C A Y
DraIII  (2550)
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.
PsiI  (2422)
1 site
T T A T A A A A T A T T
XmnI  (1955)
1 site
G A A N N N N T T C C T T N N N N A A G
BsaHI  (1893)
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  (1836)
1 site
A G T A C T T C A T G A
TatI  (1834)
1 site
W G T A C W W C A T G W
NmeAIII  (1504)
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  (1426)
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  (1417)
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.
PspOMI  (2971)
1 site
G G G C C C C C C G G G
EcoO109I  (2972)
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  (2975)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BtgI  (2977)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
NcoI  (2977)
1 site
C C A T G G G G T A C C
StyI  (2977)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
PmeI  (2986)
1 site
G T T T A A A C C A A A T T T G
EcoRI  (2991)
1 site
G A A T T C C T T A A G
BspEI  (2997)
1 site
T C C G G A A G G C C T
* Blocked by Dam methylation.
StuI  (3007)
1 site
A G G C C T T C C G G A
End  (3013)
0 sites
Start  (0)
0 sites
BamHI  (5)
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  (11)
1 site
A C T A G T T G A T C A
BfuAI  (18)
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  (18)
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.
EagI  (18)
1 site
C G G C C G G C C G G C
NotI  (18)
1 site
G C G G C C G C C G C C G G C G
PstI  (29)
1 site
C T G C A G G A C G T C
SbfI  (29)
1 site
C C T G C A G G G G A C G T C C
SalI  (31)
1 site
G T C G A C C A G C T G
AccI  (32)
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  (33)
1 site
G T Y R A C C A R Y T G
NdeI  (41)
1 site
C A T A T G G T A T A C

Prolonged incubation with NdeI may lead to removal of additional
nucleotides.
Eco53kI  (51)
1 site
G A G C T C C T C G A G
AvaI  (52)
1 site
C Y C G R G G R G C Y C

Sticky ends from different AvaI sites may not be compatible.
BsoBI  (52)
1 site
C Y C G R G G R G C Y C

Sticky ends from different BsoBI sites may not be compatible.
BsoBI is typically used at 37°C, but can be used at temperatures
up to 65°C.
PaeR7I  (52)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (52)
1 site
V C T C G A G B B G A G C T C V
XhoI  (52)
1 site
C T C G A G G A G C T C
BmeT110I  (53)
1 site
C Y C G R G G R G C Y C
SacI  (53)
1 site
G A G C T C C T C G A G
MluI  (60)
1 site
A C G C G T T G C G C A
NsiI  (73)
1 site
A T G C A T T A C G T A
BspQI  (347)
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  (347)
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.
PciI  (463)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
NspI  (467)
1 site
R C A T G Y Y G T A C R
BseYI  (767)
1 site
C C C A G C G G G T C G

After cleavage, BseYI can remain bound to DNA and alter its
electrophoretic mobility.
PspFI  (771)
1 site
C C C A G C G G G T C G
AlwNI  (879)
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.
AhdI  (1356)
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.
AmpR
1283 .. 2143  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1283 .. 2074  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1283 .. 2143  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   2075 .. 2143  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1283 .. 2143  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
524 .. 1112  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
524 .. 1112  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
2326 .. 2781  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
2326 .. 2781  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
LacZα
1 .. 126  =  126 bp
42 amino acids  =  4.6 kDa
Product: LacZα fragment of β-galactosidase
LacZα
1 .. 126  =  126 bp
42 amino acids  =  4.6 kDa
Product: LacZα fragment of β-galactosidase
AmpR promoter
2144 .. 2248  =  105 bp
AmpR promoter
2144 .. 2248  =  105 bp
MCS
2971 .. 3013  =  43 bp
multiple cloning site
MCS
2971 .. 3013  =  43 bp
multiple cloning site
lac promoter
170 .. 200  =  31 bp
   Segment 3:  -10  
   170 .. 176  =  7 bp
promoter for the E. coli lac operon
lac promoter
170 .. 200  =  31 bp
   Segment 2:  
   177 .. 194  =  18 bp
promoter for the E. coli lac operon
lac promoter
170 .. 200  =  31 bp
   Segment 1:  -35  
   195 .. 200  =  6 bp
promoter for the E. coli lac operon
lac promoter
170 .. 200  =  31 bp
3 segments
promoter for the E. coli lac operon
T7 promoter
2945 .. 2963  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
2945 .. 2963  =  19 bp
promoter for bacteriophage T7 RNA polymerase
lac operator
146 .. 162  =  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
146 .. 162  =  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).
M13 fwd
2922 .. 2938  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
2922 .. 2938  =  17 bp
common sequencing primer, one of multiple similar
variants
LacZα
2758 .. 3012  =  255 bp
84 amino acids  =  9.7 kDa
Product: LacZα fragment of β-galactosidase
LacZα
2758 .. 3012  =  255 bp
84 amino acids  =  9.7 kDa
Product: LacZα fragment of β-galactosidase
MCS
1 .. 65  =  65 bp
multiple cloning site
MCS
1 .. 65  =  65 bp
multiple cloning site
SP6 promoter
86 .. 104  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
SP6 promoter
86 .. 104  =  19 bp
promoter for bacteriophage SP6 RNA polymerase
M13 rev
122 .. 138  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
122 .. 138  =  17 bp
common sequencing primer, one of multiple similar
variants
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