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Plasmid Files

pTriEx™-4 Ek/LIC (linearized)

Linearized insect, bacterial, and mammalian vector for ligation-independent cloning (LIC), encoding an N-terminal 6xHis-S-Tag and a C-terminal HSV-8xHis.

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pTriEx-4 Ek_LIC (linearized) Sequence and MappTriEx-4 Ek_LIC (linearized).dna
Map and Sequence File   
Sequence Author:  Novagen (EMD Millipore)
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 AanI (1287) AanI (730) Bsu36I (196) AanI (3298) AanI (3387) CMV enhancer NcoI (5090) XcmI (5116) PflMI (5160) TspMI - XmaI (5192) SmaI (5194) End (5223) Start (0) EcoRV (20) Eco53kI (27) SacI (29) BamHI (33) EcoRI (39) BglII (47) AscI - BssHII (55) PstI - SbfI (65) Acc65I (73) AgeI (76) KpnI (77) BstBI (82) HindIII (85) EagI - NotI (92) PvuII (102) BstZ17I (107) PmlI (112) PaeR7I - XhoI (154) DraIII (186) pTriEx™-4 Ek/LIC 5223 bp
AanI  (1287)
4 sites
T T A T A A A A T A T T
AanI  (730)
4 sites
T T A T A A A A T A T T
Bsu36I  (196)
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.
AanI  (3298)
4 sites
T T A T A A A A T A T T
AanI  (3387)
4 sites
T T A T A A A A T A T T
NcoI  (5090)
1 site
C C A T G G G G T A C C
XcmI  (5116)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
PflMI  (5160)
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.
TspMI  (5192)
1 site
C C C G G G G G G C C C
XmaI  (5192)
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  (5194)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
End  (5223)
0 sites
Start  (0)
0 sites
EcoRV  (20)
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.
Eco53kI  (27)
1 site
G A G C T C C T C G A G
SacI  (29)
1 site
G A G C T C C T C G A G
BamHI  (33)
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  (39)
1 site
G A A T T C C T T A A G
BglII  (47)
1 site
A G A T C T T C T A G A
AscI  (55)
1 site
G G C G C G C C C C G C G C G G
BssHII  (55)
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.
PstI  (65)
1 site
C T G C A G G A C G T C
SbfI  (65)
1 site
C C T G C A G G G G A C G T C C
Acc65I  (73)
1 site
G G T A C C C C A T G G
AgeI  (76)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
KpnI  (77)
1 site
G G T A C C C C A T G G
BstBI  (82)
1 site
T T C G A A A A G C T T
HindIII  (85)
1 site
A A G C T T T T C G A A
EagI  (92)
1 site
C G G C C G G C C G G C
NotI  (92)
1 site
G C G G C C G C C G C C G G C G
PvuII  (102)
1 site
C A G C T G G T C G A C
BstZ17I  (107)
1 site
G T A T A C C A T A T G
PmlI  (112)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
PaeR7I  (154)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (154)
1 site
C T C G A G G A G C T C
DraIII  (186)
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.
AmpR
2056 .. 2916  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   2056 .. 2847  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2056 .. 2916  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   2848 .. 2916  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2056 .. 2916  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
baculovirus recombination region (lef2/ORF603)
3159 .. 3986  =  828 bp
contains ORF603 and part of lef2
baculovirus recombination region (lef2/ORF603)
3159 .. 3986  =  828 bp
contains ORF603 and part of lef2
baculovirus recombination region (ORF1629)
541 .. 1246  =  706 bp
contains part of ORF1629
baculovirus recombination region (ORF1629)
541 .. 1246  =  706 bp
contains part of ORF1629
ori
1456 .. 2043  =  588 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1456 .. 2043  =  588 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
Exon 2
4841 .. 5236  =  396 bp
Exon 2
4841 .. 5236  =  396 bp
Exon 2
1 .. 362  =  362 bp
Exon 2
1 .. 362  =  362 bp
CMV enhancer
4054 .. 4357  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
4054 .. 4357  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
4359 .. 4558  =  200 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
4359 .. 4558  =  200 bp
human cytomegalovirus (CMV) immediate early
promoter
intron
4677 .. 4840  =  164 bp
intron
4677 .. 4840  =  164 bp
Exon 1
4575 .. 4676  =  102 bp
Exon 1
4575 .. 4676  =  102 bp
T7 terminator
481 .. 528  =  48 bp
transcription terminator for bacteriophage T7 RNA
polymerase
T7 terminator
481 .. 528  =  48 bp
transcription terminator for bacteriophage T7 RNA
polymerase
ORF603
3321 .. 3926  =  606 bp
201 amino acids  =  23.6 kDa
Product: baculovirus ORF603 protein
ORF603
3321 .. 3926  =  606 bp
201 amino acids  =  23.6 kDa
Product: baculovirus ORF603 protein
ORF1629
658 .. 1246  =  589 bp
195 amino acids  =  22.3 kDa
Product: baculovirus capsid-associated protein
required for viral replication
ORF1629
658 .. 1246  =  589 bp
195 amino acids  =  22.3 kDa
Product: baculovirus capsid-associated protein
required for viral replication
lef2
3159 .. 3283  =  125 bp
40 amino acids  =  4.5 kDa
Product: baculovirus late expression factor 2
lef2
3159 .. 3283  =  125 bp
40 amino acids  =  4.5 kDa
Product: baculovirus late expression factor 2
rabbit globin terminator
360 .. 471  =  112 bp
rabbit globin terminator
360 .. 471  =  112 bp
p10 promoter
4968 .. 5077  =  110 bp
baculovirus promoter for expression in insect cells
p10 promoter
4968 .. 5077  =  110 bp
baculovirus promoter for expression in insect cells
S-Tag
5125 .. 5169  =  45 bp
15 amino acids  =  1.7 kDa
Product: affinity and epitope tag derived from
pancreatic ribonuclease A
S-Tag
5125 .. 5169  =  45 bp
15 amino acids  =  1.7 kDa
Product: affinity and epitope tag derived from
pancreatic ribonuclease A
HSV tag
121 .. 153  =  33 bp
11 amino acids  =  1.2 kDa
Product: HSV (herpes simplex virus) epitope tag
HSV tag
121 .. 153  =  33 bp
11 amino acids  =  1.2 kDa
Product: HSV (herpes simplex virus) epitope tag
lac operator
4928 .. 4952  =  25 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
4928 .. 4952  =  25 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).
8xHis
160 .. 183  =  24 bp
8 amino acids  =  1.1 kDa
Product: 8xHis affinity tag
8xHis
160 .. 183  =  24 bp
8 amino acids  =  1.1 kDa
Product: 8xHis affinity tag
T7 promoter
4909 .. 4927  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
4909 .. 4927  =  19 bp
promoter for bacteriophage T7 RNA polymerase
6xHis
5098 .. 5115  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
5098 .. 5115  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
thrombin site
5185 .. 5202  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
thrombin site
5185 .. 5202  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
enterokinase site
5221 .. 5235  =  15 bp
5 amino acids  =  606.5 Da
Product: enterokinase recognition and cleavage site
enterokinase site
5221 .. 5235  =  15 bp
5 amino acids  =  606.5 Da
Product: enterokinase recognition and cleavage site
poly(A) signal
338 .. 343  =  6 bp
poly(A) signal
338 .. 343  =  6 bp
RBS
5079 .. 5084  =  6 bp
ribosome binding site
RBS
5079 .. 5084  =  6 bp
ribosome binding site
ATG
5092 .. 5094  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
5092 .. 5094  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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