Basic Vector Information
pET-29b(+) 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 ℃.
pET-29b(+) vector Sequence
LOCUS pET-29b(+). 5370 bp DNA circular SYN 01-JAN-1980 DEFINITION Bacterial vector for expression of N-terminally S-tagged proteins with a thrombin site. For other reading frames, use pET-29a(+) or pET-29c(+). ACCESSION . VERSION . KEYWORDS pET-29b(+). SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5370) AUTHORS Novagen (EMD Millipore) TITLE Direct Submission REFERENCE 2 (bases 1 to 5370) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article" FEATURES Location/Qualifiers source 1..5370 /mol_type="other DNA" /organism="synthetic DNA construct" terminator complement(26..73) /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" CDS complement(140..157) /label=6xHis /note="6xHis affinity tag" misc_feature 158..203 /label=MCS /note="MCS" /note="multiple cloning site" CDS complement(215..232) /label=thrombin site /note="thrombin recognition and cleavage site" CDS complement(248..292) /label=S-Tag /note="affinity and epitope tag derived from pancreatic ribonuclease A" CDS complement(293..295) /codon_start=1 /product="start codon" /label=start codon /note="ATG" /translation="M" RBS complement(303..325) /label=RBS /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" protein_bind complement(340..364) /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)." promoter complement(365..383) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" promoter 696..773 /label=lacI promoter CDS 774..1853 /label=lacI /note="lac repressor" protein_bind 1869..1890 /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." CDS 2665..2853 /label=rop /note="Rop protein, which maintains plasmids at low copy number" misc_feature 2958..3100 /label=bom /note="basis of mobility region from pBR322" rep_origin complement(3286..3874) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS 3996..4808 /label=KanR /note="aminoglycoside phosphotransferase" rep_origin complement(4904..5359) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis"
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