Basic Vector Information
- Vector Name:
- pMCSG23
- Antibiotic Resistance:
- Streptomycin
- Length:
- 4971 bp
- Type:
- Structural Genomics Vectors
- Replication origin:
- CloDF13 ori
- Source/Author:
- Eschenfeldt WH, Lucy S, Millard CS, Joachimiak A, Mark ID.
- Copy Number:
- High copy number
pMCSG23 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 ℃.
pMCSG23 vector Sequence
LOCUS pMCSG23. 4971 bp DNA circular SYN 01-JAN-1980
DEFINITION Bacterial coexpression vector with a 6xHis-MBP-TEV leader and a
CloDF13 origin, for high-throughput purification of recombinant
proteins.
ACCESSION .
VERSION .
KEYWORDS pMCSG23.
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4971)
AUTHORS Eschenfeldt WH, Lucy S, Millard CS, Joachimiak A, Mark ID.
TITLE A family of LIC vectors for high-throughput cloning and purification
of proteins.
JOURNAL Methods Mol. Biol. 2009;498:105-15.
PUBMED 18988021
REFERENCE 2 (bases 1 to 4971)
AUTHORS Midwest Center for Structural Genomics
TITLE Direct Submission
REFERENCE 3 (bases 1 to 4971)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Methods
Mol. Biol."; date: "2009"; volume: "498"; pages: "105-15"
COMMENT SGRef: number: 2; 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..4971
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind 3..27
/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 42..64
/label=RBS
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 71..73
/codon_start=1
/product="start codon"
/label=start codon
/note="ATG"
/translation="M"
CDS 83..100
/label=6xHis
/note="6xHis affinity tag"
promoter 214..232
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 233..257
/label=lac repressor encoded by lacI binding site
/bound_moiety="lac repressor encoded by lacI"
/note="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 286..291
/note="ribosome binding site"
CDS 300..302
/codon_start=1
/product="start codon"
/label=start codon
/note="ATG"
/translation="M"
CDS 303..320
/label=6xHis
/note="6xHis affinity tag"
CDS 345..1442
/label=MBP
/note="maltose binding protein from E. coli"
CDS 1464..1484
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
CDS 1556..1600
/label=S-Tag
/note="affinity and epitope tag derived from pancreatic
ribonuclease A"
terminator 1652..1699
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
CDS complement(1873..2661)
/label=SmR
/note="aminoglycoside adenylyltransferase (Murphy, 1985)"
promoter complement(2662..2753)
/label=AmpR promoter
rep_origin complement(2801..3539)
/direction=LEFT
/label=CloDF13 ori
/note="Plasmids containing the CloDF13 (CDF) origin of
replication can be propagated in E. coli cells that contain
additional plasmids with compatible origins."
protein_bind complement(3715..3736)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(3752..4831)
/label=lacI
/note="lac repressor"
promoter complement(4832..4909)
/label=lacI promoter
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