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Plasmid Files

pCoBlast

Selection plasmid for insect cells, with a blasticidin resistance gene under control of the copia promoter.

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pCoBlast Sequence and MappCoBlast.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Sciences)
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 AatII (3842) ZraI (3840) SspI (3724) XmnI (3519) ScaI (3400) BpmI (2990) BsaI (2981) AhdI (2920) AlwNI (2443) AflIII - PciI (2027) PfoI (46) NdeI (184) BstAPI (185) KasI (235) NarI (236) SfoI (237) PluTI (239) HindIII (399) BfuAI - BspMI (404) SphI (409) PstI - SbfI (415) SalI (417) AccI (418) HincII (419) XbaI (423) BamHI (429) BbsI (882) EagI (913) PpuMI (956) NruI * (1028) BsaBI * (1095) BbvCI - Bpu10I (1251) BlpI (1337) PasI (1408) AleI (1437) AbsI - PaeR7I - PspXI - XhoI (1505) BsgI (1514) NheI (1515) BmtI (1519) BsmI (1604) Acc65I (1660) KpnI (1664) Eco53kI (1668) SacI (1670) EcoRI (1672) BspQI - SapI (1911) pCoBlast 3907 bp
AatII  (3842)
1 site
G A C G T C C T G C A G
ZraI  (3840)
1 site
G A C G T C C T G C A G
SspI  (3724)
1 site
A A T A T T T T A T A A
XmnI  (3519)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (3400)
1 site
A G T A C T T C A T G A
BpmI  (2990)
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  (2981)
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  (2920)
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  (2443)
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.
AflIII  (2027)
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  (2027)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PfoI  (46)
1 site
T C C N G G A A G G N C C T

Sticky ends from different PfoI sites may not be compatible.
NdeI  (184)
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.
BstAPI  (185)
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.
KasI  (235)
1 site
G G C G C C C C G C G G
NarI  (236)
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  (237)
1 site
G G C G C C C C G C G G
PluTI  (239)
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.
HindIII  (399)
1 site
A A G C T T T T C G A A
BfuAI  (404)
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  (404)
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.
SphI  (409)
1 site
G C A T G C C G T A C G
PstI  (415)
1 site
C T G C A G G A C G T C
SbfI  (415)
1 site
C C T G C A G G G G A C G T C C
SalI  (417)
1 site
G T C G A C C A G C T G
AccI  (418)
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  (419)
1 site
G T Y R A C C A R Y T G
XbaI  (423)
1 site
T C T A G A A G A T C T
BamHI  (429)
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.
BbsI  (882)
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.
EagI  (913)
1 site
C G G C C G G C C G G C
PpuMI  (956)
1 site
R G G W C C Y Y C C W G G R

Sticky ends from different PpuMI sites may not be compatible.
NruI  (1028)
1 site
T C G C G A A G C G C T
* Blocked by Dam methylation.
BsaBI  (1095)
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.
BbvCI  (1251)
1 site
C C T C A G C G G A G T C G
Bpu10I  (1251)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
This recognition sequence is asymmetric, so ligating sticky ends
generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
BlpI  (1337)
1 site
G C T N A G C C G A N T C G

Sticky ends from different BlpI sites may not be compatible.
PasI  (1408)
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.
AleI  (1437)
1 site
C A C N N N N G T G G T G N N N N C A C
AbsI  (1505)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (1505)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (1505)
1 site
V C T C G A G B B G A G C T C V
XhoI  (1505)
1 site
C T C G A G G A G C T C
BsgI  (1514)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14

Efficient cleavage requires at least two copies of the BsgI
recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
NheI  (1515)
1 site
G C T A G C C G A T C G
BmtI  (1519)
1 site
G C T A G C C G A T C G
BsmI  (1604)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
Acc65I  (1660)
1 site
G G T A C C C C A T G G
KpnI  (1664)
1 site
G G T A C C C C A T G G
Eco53kI  (1668)
1 site
G A G C T C C T C G A G
SacI  (1670)
1 site
G A G C T C C T C G A G
EcoRI  (1672)
1 site
G A A T T C C T T A A G
BspQI  (1911)
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  (1911)
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
2847 .. 3707  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   2847 .. 3638  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2847 .. 3707  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   3639 .. 3707  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2847 .. 3707  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
2088 .. 2676  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2088 .. 2676  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
BSD
797 .. 1195  =  399 bp
132 amino acids  =  13.7 kDa
Product: blasticidin S deaminase
confers resistance to blasticidin
BSD
797 .. 1195  =  399 bp
132 amino acids  =  13.7 kDa
Product: blasticidin S deaminase
confers resistance to blasticidin
copia promoter
501 .. 780  =  280 bp
strong promoter from the Drosophila transposable
element copia (Sinclair et al., 1986)
copia promoter
501 .. 780  =  280 bp
strong promoter from the Drosophila transposable
element copia (Sinclair et al., 1986)
SV40 poly(A) signal
1524 .. 1658  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1524 .. 1658  =  135 bp
SV40 polyadenylation signal
AmpR promoter
3708 .. 3812  =  105 bp
AmpR promoter
3708 .. 3812  =  105 bp
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