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Plasmid Files

pNIC-CH (linearized)

Linearized bacterial vector for ligation-independent cloning (LIC), encoding a C-terminal 6xHis tag.

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pNIC-CH (linearized) Sequence and MappNIC-CH (linearized).dna
Map and Sequence File   
Sequence Author:  Structural Genomics Consortium
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 BstAPI (4790) MluI (4466) BclI * (4452) BstEII (4284) NmeAIII (4266) ApaI (4263) PspOMI (4259) BssHII (4055) EcoRV (4020) HpaI (3964) PshAI (3625) BglI (3409) FspI - FspAI (3388) PpuMI (3360) PflFI - Tth111I (2623) BstZ17I (2598) TatI (2561) BspQI - SapI (2482) PciI (2365) BssS α I (2192) SphI (4999) SgrAI (5147) BglII (5188) T7 promoter XbaI (5254) End (5276) RBS Start (0) BamHI (22) EcoRI (28) Eco53kI (36) SacI (38) SalI (41) HindIII (47) EagI - NotI (54) PaeR7I - PspXI - XhoI (62) BlpI (141) StyI (163) DraIII (469) PsiI (594) AsiSI - PvuI (1169) TspMI - XmaI (1291) SmaI (1293) BspDI - ClaI (1474) NruI (1510) AcuI (1823) AlwNI (1956) pNIC-CH 5276 bp
BstAPI  (4790)
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.
MluI  (4466)
1 site
A C G C G T T G C G C A
BclI  (4452)
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.
BstEII  (4284)
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.
NmeAIII  (4266)
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).
ApaI  (4263)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PspOMI  (4259)
1 site
G G G C C C C C C G G G
BssHII  (4055)
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.
EcoRV  (4020)
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.
HpaI  (3964)
1 site
G T T A A C C A A T T G
PshAI  (3625)
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.
BglI  (3409)
1 site
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
FspI  (3388)
1 site
T G C G C A A C G C G T
FspAI  (3388)
1 site
R T G C G C A Y Y A C G C G T R
PpuMI  (3360)
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.
PflFI  (2623)
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  (2623)
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.
BstZ17I  (2598)
1 site
G T A T A C C A T A T G
TatI  (2561)
1 site
W G T A C W W C A T G W
BspQI  (2482)
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  (2482)
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  (2365)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BssSαI  (2192)
1 site
C A C G A G G T G C T C
SphI  (4999)
1 site
G C A T G C C G T A C G
SgrAI  (5147)
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.
BglII  (5188)
1 site
A G A T C T T C T A G A
XbaI  (5254)
1 site
T C T A G A A G A T C T
End  (5276)
0 sites
Start  (0)
0 sites
BamHI  (22)
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  (28)
1 site
G A A T T C C T T A A G
Eco53kI  (36)
1 site
G A G C T C C T C G A G
SacI  (38)
1 site
G A G C T C C T C G A G
SalI  (41)
1 site
G T C G A C C A G C T G
HindIII  (47)
1 site
A A G C T T T T C G A A
EagI  (54)
1 site
C G G C C G G C C G G C
NotI  (54)
1 site
G C G G C C G C C G C C G G C G
PaeR7I  (62)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (62)
1 site
V C T C G A G B B G A G C T C V
XhoI  (62)
1 site
C T C G A G G A G C T C
BlpI  (141)
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.
StyI  (163)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
DraIII  (469)
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  (594)
1 site
T T A T A A A A T A T T
AsiSI  (1169)
1 site
G C G A T C G C C G C T A G C G
PvuI  (1169)
1 site
C G A T C G G C T A G C
TspMI  (1291)
1 site
C C C G G G G G G C C C
XmaI  (1291)
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  (1293)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
BspDI  (1474)
1 site
A T C G A T T A G C T A
ClaI  (1474)
1 site
A T C G A T T A G C T A
NruI  (1510)
1 site
T C G C G A A G C G C T
AcuI  (1823)
1 site
C T G A A G ( N ) 14 N N G A C T T C ( N ) 14

Efficient cleavage requires at least two copies of the AcuI
recognition sequence.
Sticky ends from different AcuI sites may not be compatible.
After cleavage, AcuI can remain bound to DNA and alter its
electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
AlwNI  (1956)
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.
lacI
3739 .. 4821  =  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
3739 .. 4821  =  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).
KanR
784 .. 1599  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
KanR
784 .. 1599  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
ori
1721 .. 2309  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1721 .. 2309  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
236 .. 691  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
236 .. 691  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
rop
2739 .. 2930  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at
low copy number
rop
2739 .. 2930  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at
low copy number
lacI promoter
4822 .. 4899  =  78 bp
lacI promoter
4822 .. 4899  =  78 bp
T7 terminator
152 .. 199  =  48 bp
transcription terminator for bacteriophage T7 RNA
polymerase
T7 terminator
152 .. 199  =  48 bp
transcription terminator for bacteriophage T7 RNA
polymerase
lac operator
5227 .. 5251  =  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
5227 .. 5251  =  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).
T7 promoter
5208 .. 5226  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
5208 .. 5226  =  19 bp
promoter for bacteriophage T7 RNA polymerase
6xHis
1 .. 18  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
1 .. 18  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
68 .. 85  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
68 .. 85  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
RBS
5282 .. 5287  =  6 bp
ribosome binding site
RBS
5282 .. 5287  =  6 bp
ribosome binding site
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