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Plasmid Files

pGEX-5X-1

Bacterial vector for expressing GST fusion proteins with a Factor Xa site. For other reading frames, use pGEX‑5X-2 or pGEX‑5X-3.

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pGEX-5X-1.dna
Map and Sequence File:    Download    Open   
Sequence Author:  GE Healthcare
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BtgI (4872) Bsu36I (4764) PluTI (4314) SfoI (4312) NarI * (4311) KasI (4310) HpaI (4177) EcoRV (4121) BssHII (4082) ApaI - BanII (3882) PspOMI (3878) BstEII (3852) MluI (3671) BstAPI (3354) PflMI (3253) AlwNI (2646) BfuAI - BspMI (62) EcoNI (268) MscI (465) BstBI (655) SwaI (685) pGEX 5' Sequencing Primer (869 .. 891) BamHI (934) EcoRI (942) TspMI - XmaI (947) SmaI (949) SalI (952) AccI (953) PaeR7I - PspXI - XhoI (957) EagI - NotI (963) PfoI (1039) pGEX 3' Sequencing Primer (1022 .. 1044) PflFI - Tth111I (1142) BsaAI (1149) ZraI (1246) AatII (1248) PstI (1925) BsaI (2101) AhdI (2167) pGEX-5X-1 4972 bp
BtgI  (4872)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
Bsu36I  (4764)
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.
PluTI  (4314)
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.
SfoI  (4312)
1 site
G G C G C C C C G C G G
NarI  (4311)
1 site
G G C G C C C C G C G G
* Blocked by Dcm methylation.
Efficient cleavage requires at least two copies of the NarI recognition sequence.
KasI  (4310)
1 site
G G C G C C C C G C G G
HpaI  (4177)
1 site
G T T A A C C A A T T G
EcoRV  (4121)
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.
BssHII  (4082)
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.
ApaI  (3882)
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  (3882)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
PspOMI  (3878)
1 site
G G G C C C C C C G G G
BstEII  (3852)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
MluI  (3671)
1 site
A C G C G T T G C G C A
BstAPI  (3354)
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.
PflMI  (3253)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
AlwNI  (2646)
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.
BfuAI  (62)
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  (62)
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.
EcoNI  (268)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
MscI  (465)
1 site
T G G C C A A C C G G T
BstBI  (655)
1 site
T T C G A A A A G C T T
SwaI  (685)
1 site
A T T T A A A T T A A A T T T A

SwaI is typically used at 25°C, but is 50% active at 37°C.
BamHI  (934)
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.
EcoRI  (942)
1 site
G A A T T C C T T A A G
TspMI  (947)
1 site
C C C G G G G G G C C C
XmaI  (947)
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.
SmaI  (949)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
SalI  (952)
1 site
G T C G A C C A G C T G
AccI  (953)
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.
PaeR7I  (957)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (957)
1 site
V C T C G A G B B G A G C T C V
XhoI  (957)
1 site
C T C G A G G A G C T C
EagI  (963)
1 site
C G G C C G G C C G G C
NotI  (963)
1 site
G C G G C C G C C G C C G G C G
PfoI  (1039)
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.
PflFI  (1142)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (1142)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
BsaAI  (1149)
1 site
Y A C G T R R T G C A Y
ZraI  (1246)
1 site
G A C G T C C T G C A G
AatII  (1248)
1 site
G A C G T C C T G C A G
PstI  (1925)
1 site
C T G C A G G A C G T C
BsaI  (2101)
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  (2167)
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.
pGEX 5' Sequencing Primer
23-mer  /  65% GC
1 binding site
869 .. 891  =  23 annealed bases
Tm  =  67°C
pGEX 3' Sequencing Primer
23-mer  /  65% GC
1 binding site
1022 .. 1044  =  23 annealed bases
Tm  =  66°C
lacI
3321 .. 4403  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
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).
lacI
3321 .. 4403  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
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).
AmpR
1380 .. 2240  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   1380 .. 1448  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1380 .. 2240  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   1449 .. 2240  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1380 .. 2240  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
GST
258 .. 911  =  654 bp
218 amino acids  =  25.5 kDa
Product: glutathione S-transferase from Schistosoma japonicum
GST
258 .. 911  =  654 bp
218 amino acids  =  25.5 kDa
Product: glutathione S-transferase from Schistosoma japonicum
ori
2411 .. 2999  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2411 .. 2999  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
AmpR promoter
1275 .. 1379  =  105 bp
AmpR promoter
1275 .. 1379  =  105 bp
lacIq promoter
3243 .. 3320  =  78 bp
In the lacIq allele, a single base change in the promoter boosts expression of the lacI gene about 10-fold.
lacIq promoter
3243 .. 3320  =  78 bp
In the lacIq allele, a single base change in the promoter boosts expression of the lacI gene about 10-fold.
MCS
934 .. 969  =  36 bp
multiple cloning site
MCS
934 .. 969  =  36 bp
multiple cloning site
tac promoter
183 .. 211  =  29 bp
   Segment 1:  -35  
   183 .. 188  =  6 bp
strong E. coli promoter; hybrid between the trp and lac UV5 promoters
tac promoter
183 .. 211  =  29 bp
   Segment 2:  
   189 .. 204  =  16 bp
strong E. coli promoter; hybrid between the trp and lac UV5 promoters
tac promoter
183 .. 211  =  29 bp
   Segment 3:  -10  
   205 .. 211  =  7 bp
strong E. coli promoter; hybrid between the trp and lac UV5 promoters
tac promoter
183 .. 211  =  29 bp
3 segments
strong E. coli promoter; hybrid between the trp and lac UV5 promoters
lac operator
219 .. 235  =  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
219 .. 235  =  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).
Factor Xa site
921 .. 932  =  12 bp
4 amino acids  =  473.5 Da
Product: Factor Xa recognition and cleavage site
Factor Xa site
921 .. 932  =  12 bp
4 amino acids  =  473.5 Da
Product: Factor Xa recognition and cleavage site
stop codons
974 .. 984  =  11 bp
stop codons in all three reading frames
stop codons
974 .. 984  =  11 bp
stop codons in all three reading frames
ORF:  3254 .. 3571  =  318 bp
ORF:  105 amino acids  =  11.2 kDa
ORF:  4160 .. 4423  =  264 bp
ORF:  87 amino acids  =  8.9 kDa
ORF:  4526 .. 81  =  528 bp
ORF:  175 amino acids  =  19.7 kDa
ORF:  258 .. 980  =  723 bp
ORF:  240 amino acids  =  28.0 kDa
ORF:  1380 .. 2240  =  861 bp
ORF:  286 amino acids  =  31.5 kDa
ORF:  3444 .. 4403  =  960 bp
ORF:  319 amino acids  =  34.1 kDa
ORF:  4911 .. 178  =  240 bp
ORF:  79 amino acids  =  9.0 kDa
ORF:  695 .. 1033  =  339 bp
ORF:  112 amino acids  =  12.4 kDa
ORF:  1844 .. 2110  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  4186 .. 4437  =  252 bp
ORF:  83 amino acids  =  9.1 kDa
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