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pGAPZ A

Pichia pastoris vector for constitutive intracellular expression of a protein. For other reading frames, use pGAPZ B or pGAPZ C.

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pGAPZ A.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Thermo Fisher (Invitrogen)
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NsiI (2877) AlwNI (2567) BciVI (2354) BssS α I (2324) PciI (2151) MluI (2062) BsrGI (2055) AclI (2021) EcoRV (1888) StuI * (1854) KflI (1846) DraIII (1803) FseI (1726) SexAI * (1613) SgrAI (1564) SmaI (1540) TspMI - XmaI (1538) AatII (1496) ZraI (1494) BssHII - MauBI (1486) BglII (1) TsoI (61) BfuAI - BspMI (96) AvrII (190) BsrDI (246) BspQI - SapI (251) PasI (301) PshAI (351) BspHI (355) DraI (439) MfeI (469) BstBI (485) EcoRI (495) BsaAI - PmlI (502) SfiI (512) Acc65I (528) KpnI (532) PaeR7I - PspXI - XhoI (533) SacII (542) NotI (545) PspOMI (558) ApaI - BanII (562) SalI (603) AccI (604) AgeI (684) BsmI (725) BsiWI (816) BclI * (832) PvuII (850) BamHI (965) BtgZI (1024) AseI (1392) NcoI (1448) MscI (1453) pGAPZ A 2884 bp
NsiI  (2877)
1 site
A T G C A T T A C G T A
AlwNI  (2567)
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.
BciVI  (2354)
1 site
G T A T C C ( N ) 5 N C A T A G G ( N ) 5

The 1-base overhangs produced by BciVI may be hard to ligate.
Sticky ends from different BciVI sites may not be compatible.
BssSαI  (2324)
1 site
C A C G A G G T G C T C
PciI  (2151)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
MluI  (2062)
1 site
A C G C G T T G C G C A
BsrGI  (2055)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
AclI  (2021)
1 site
A A C G T T T T G C A A
EcoRV  (1888)
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.
StuI  (1854)
1 site
A G G C C T T C C G G A
* Blocked by Dcm methylation.
KflI  (1846)
1 site
G G G W C C C C C C W G G G

Sticky ends from different KflI sites may not be compatible.
DraIII  (1803)
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.
FseI  (1726)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
SexAI  (1613)
1 site
A C C W G G T T G G W C C A
* Blocked by Dcm methylation.
Sticky ends from different SexAI sites may not be compatible.
SgrAI  (1564)
1 site
C R C C G G Y G G Y G G C C R C

Efficient cleavage requires at least two copies of the SgrAI recognition sequence.
SmaI  (1540)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
TspMI  (1538)
1 site
C C C G G G G G G C C C
XmaI  (1538)
1 site
C C C G G G G G G C C C

Efficient cleavage requires at least two copies of the XmaI recognition sequence.
Full cleavage with XmaI may require a long incubation.
AatII  (1496)
1 site
G A C G T C C T G C A G
ZraI  (1494)
1 site
G A C G T C C T G C A G
BssHII  (1486)
1 site
G C G C G C C G C G C G

BssHII is typically used at 50°C, but is 75% active at 37°C.
MauBI  (1486)
1 site
C G C G C G C G G C G C G C G C
BglII  (1)
1 site
A G A T C T T C T A G A
TsoI  (61)
1 site
T A R C C A ( N ) 9 N N A T Y G G T ( N ) 9

Sticky ends from different TsoI sites may not be compatible.
After cleavage, TsoI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
BfuAI  (96)
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  (96)
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.
AvrII  (190)
1 site
C C T A G G G G A T C C
BsrDI  (246)
1 site
G C A A T G N N C G T T A C

Sticky ends from different BsrDI sites may not be compatible.
BspQI  (251)
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  (251)
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.
PasI  (301)
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.
PshAI  (351)
1 site
G A C N N N N G T C C T G N N N N C A G

PshAI quickly loses activity at 37°C, but can be used at 25°C for long incubations.
BspHI  (355)
1 site
T C A T G A A G T A C T
DraI  (439)
1 site
T T T A A A A A A T T T
MfeI  (469)
1 site
C A A T T G G T T A A C
BstBI  (485)
1 site
T T C G A A A A G C T T
EcoRI  (495)
1 site
G A A T T C C T T A A G
BsaAI  (502)
1 site
Y A C G T R R T G C A Y
PmlI  (502)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
SfiI  (512)
1 site
G G C C N N N N N G G C C C C G G N N N N N C C G G

Efficient cleavage requires at least two copies of the SfiI recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
Acc65I  (528)
1 site
G G T A C C C C A T G G
KpnI  (532)
1 site
G G T A C C C C A T G G
PaeR7I  (533)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (533)
1 site
V C T C G A G B B G A G C T C V
XhoI  (533)
1 site
C T C G A G G A G C T C
SacII  (542)
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.
NotI  (545)
1 site
G C G G C C G C C G C C G G C G
PspOMI  (558)
1 site
G G G C C C C C C G G G
ApaI  (562)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BanII  (562)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
SalI  (603)
1 site
G T C G A C C A G C T G
AccI  (604)
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.
AgeI  (684)
1 site
A C C G G T T G G C C A
BsmI  (725)
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.
BsiWI  (816)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
BclI  (832)
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.
PvuII  (850)
1 site
C A G C T G G T C G A C
BamHI  (965)
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.
BtgZI  (1024)
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.
AseI  (1392)
1 site
A T T A A T T A A T T A
NcoI  (1448)
1 site
C C A T G G G G T A C C
MscI  (1453)
1 site
T G G C C A A C C G G T
ori
2212 .. 2800  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2212 .. 2800  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
GAP promoter
7 .. 483  =  477 bp
promoter for Pichia pastoris glyceraldehyde-3-phosphate dehydrogenase
GAP promoter
7 .. 483  =  477 bp
promoter for Pichia pastoris glyceraldehyde-3-phosphate dehydrogenase
TEF1 promoter
966 .. 1377  =  412 bp
promoter for EF-1α
TEF1 promoter
966 .. 1377  =  412 bp
promoter for EF-1α
BleoR
1450 .. 1824  =  375 bp
124 amino acids  =  13.8 kDa
Product: antibiotic-binding protein
confers resistance to bleomycin, phleomycin, and Zeocin™
BleoR
1450 .. 1824  =  375 bp
124 amino acids  =  13.8 kDa
Product: antibiotic-binding protein
confers resistance to bleomycin, phleomycin, and Zeocin™
CYC1 terminator
1890 .. 2137  =  248 bp
transcription terminator for CYC1
CYC1 terminator
1890 .. 2137  =  248 bp
transcription terminator for CYC1
AOX1 terminator
705 .. 951  =  247 bp
transcription terminator for AOX1
AOX1 terminator
705 .. 951  =  247 bp
transcription terminator for AOX1
MCS
484 .. 563  =  80 bp
multiple cloning site
MCS
484 .. 563  =  80 bp
multiple cloning site
EM7 promoter
1384 .. 1431  =  48 bp
synthetic bacterial promoter
EM7 promoter
1384 .. 1431  =  48 bp
synthetic bacterial promoter
Myc
564 .. 593  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
Myc
564 .. 593  =  30 bp
10 amino acids  =  1.2 kDa
Product: Myc (human c-Myc oncogene) epitope tag
6xHis
609 .. 626  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
609 .. 626  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
ORF:  136 .. 360  =  225 bp
ORF:  74 amino acids  =  8.0 kDa
ORF:  1450 .. 1824  =  375 bp
ORF:  124 amino acids  =  13.8 kDa
ORF:  357 .. 629  =  273 bp
ORF:  90 amino acids  =  10.6 kDa
ORF:  1396 .. 1893  =  498 bp
ORF:  165 amino acids  =  17.5 kDa
ORF:  1443 .. 1730  =  288 bp
ORF:  95 amino acids  =  10.1 kDa
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