Gene:
Gene Symbol:
No insert
Gene Name:
No insert
Original Clone ID:
None
Type:
No insert
Vector Name:
pMCSG18
Source:
M. Donnelly
Description:
Screening vector with T7 promoter, adds N-terminal His tag, TEV, LICs, GFP; amp resistance; ligation independent cloning (LIC).
Comments:
Standard LIC cloning (linearize the vector with SspI, then treat insert and vector with T4 polymerase), using these sequences for the PCR primers:
Forward: 5'-TACTTCCAATCCAATGCC-3' + GOI sequence
Reverse: 5'-TAATCCACTTCCAATAGC-3' +GOI sequence
These primers leave an alanine scar on both the N and C terminus of the GOI resulting in the following protein sequence:
His6-TEV(cleavage site)-Alanine-GOI-Alanine-GFP
Publications:
None
Authors:
M.Donnelly
Midwest Center for Structural Genomics
PSI
Midwest Center for Structural Genomics
Midwest Center for Structural Genomics
PSI
Midwest Center for Structural Genomics
EvNO00084264
Price:
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AVAILABLE
No restriction
Special MTA: PSI
Special MTA: PSI
Recommended Growth Condition:
Distributed in bacterial strain:
DH5-alpha T1 phage resistant
Antibiotic Selection:
Host Type:
bacterial
Marker:
ampicillin
Bacterial Selection Condition:
100 ug/mL ampicillin
Growth Condition:
Growth with the single antibiotic in LB at 37 degrees is recommended.
Comments:
Commonly used conditions for ampicillin resistant plasmid clones.
Vector Name:
pMCSG18
Synonyms:
pMCSG-18
Sequencing Primer:
Forward:
T7
Reverse:
T7 terminator
Description:
Screening vector with T7 promoter, adds N-terminal His tag, TEV, LICs, GFP; amp resistance; ligation independent cloning (LIC).
Comments:
pMCSG7 derivative
Size (bp):
6009
Parent Vector:
None
Properties:
bacterial expression, in vitro transcription, ligation independent cloning (LIC), with tag/fusion/marker
Author Name:
Midwest Center for Structural Genomics
Publications:
None
Vector Map:
Vector Sequence:
Sequence:
Vector Features:
Type
Name
Description
Start
End
MCS
LIC
Ligation independent cloning (LIC) site
927
958
bacterial origin
ori
ColE1-type bacterial origin of replication
3984
4666
promoter
T7
T7 promoter
1083
1099
protease cleavage site
TEV
TEV cleavage site cds
946
966
repressor binding site
lac operator
LacO fragment
1058
1080
repressor protein gene
LacI
LacI coding sequence
1613
2572
selectable marker
AmpR
ampicillin resistance gene
4761
5621
tag
His 2
C-terminal 6xHis tag
140
157
tag
GFP
GFP tag
208
927
tag
His
N-terminal 6xHis tag
991
1008
trxn termination sequence
T7 term
T7 terminator sequence
26
72
