Basic Vector Information
- Vector Name:
- pMCSG21
- Antibiotic Resistance:
- Streptomycin
- Length:
- 3852 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
pMCSG21 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 ℃.
pMCSG21 vector Sequence
LOCUS pMCSG21. 3852 bp DNA circular SYN 01-JAN-1980 DEFINITION Bacterial coexpression vector with a 6xHis-TEV leader and a CloDF13 origin, for high-throughput purification of recombinant proteins. ACCESSION . VERSION . KEYWORDS pMCSG21. SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3852) 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 3852) AUTHORS Midwest Center for Structural Genomics TITLE Direct Submission REFERENCE 3 (bases 1 to 3852) 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..3852 /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..365 /label=TEV site /note="tobacco etch virus (TEV) protease recognition and cleavage site" CDS 437..481 /label=S-Tag /note="affinity and epitope tag derived from pancreatic ribonuclease A" terminator 533..580 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" CDS complement(754..1542) /label=SmR /note="aminoglycoside adenylyltransferase (Murphy, 1985)" promoter complement(1543..1634) /label=AmpR promoter rep_origin complement(1682..2420) /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(2596..2617) /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." CDS complement(2633..3712) /label=lacI /note="lac repressor" promoter complement(3713..3790) /label=lacI promoter
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