Basic Vector Information
- Vector Name:
- pMCSG31
- Antibiotic Resistance:
- Ampicillin
- Length:
- 5822 bp
- Type:
- Structural Genomics Vectors
- Replication origin:
- ori
- Source/Author:
- Midwest Center for Structural Genomics
- Copy Number:
- High copy number
- Promoter:
- tetR/tetAs
pMCSG31 vector Vector Map
Plasmid Resuspension Protocol:
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5.Store the plasmid at -20 ℃.
pMCSG31 vector Sequence
LOCUS pMCSG31. 5822 bp DNA circular SYN 01-JAN-1980
DEFINITION Bacterial expression vector with an MBP-TVMV-6xHis-TEV leader and a
Tet-inducible TVMV protease cassette.
ACCESSION .
VERSION .
KEYWORDS pMCSG31.
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5822)
AUTHORS Midwest Center for Structural Genomics
TITLE Direct Submission
REFERENCE 2 (bases 1 to 5822)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
COMMENT For ligation-independent cloning (LIC), linearize with SspI and
treat with T4 DNA polymerase plus dGTP.
FEATURES Location/Qualifiers
source 1..5822
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 36..91
/label=tetR/tetA promoters
/note="overlapping promoters for bacterial tetR and tetA"
RBS 130..152
/label=RBS
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 184..891
/label=TVMV protease
/note="tobacco vein mottling virus NIa protease
(Nallamsetty et al., 2004)"
terminator 927..1013
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
promoter 1244..1262
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 1263..1287
/label=lac operator
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
RBS 1318..1323
/note="ribosome binding site"
CDS 1332..2432
/label=MBP
/note="maltose binding protein from E. coli"
protein_bind 2442..2466
/label=attB1
/note="recombination site for the Gateway(R) BP reaction"
CDS 2469..2489
/label=TVMV site
/note="tobacco vein mottling virus (TVMV) NIa protease
recognition and cleavage site"
CDS 2490..2507
/label=6xHis
/note="6xHis affinity tag"
CDS 2532..2552
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
CDS 2618..2635
/label=6xHis
/note="6xHis affinity tag"
terminator 2702..2749
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
CDS 2818..2841
/label=Strep-Tag II
/note="peptide that binds Strep-Tactin(R), an engineered
form
of streptavidin"
rep_origin 2932..3390
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 3416..3519
/label=AmpR promoter
CDS 3520..4377
/label=AmpR
/note="beta-lactamase"
CDS 4387..5010
/label=TetR
/note="tetracycline repressor TetR"
rep_origin 5166..5754
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
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