Basic Vector Information
- Vector Name:
- pET-28
- Antibiotic Resistance:
- Kanamycin
- Length:
- 5331 bp
- Type:
- Expression vector
- Replication origin:
- ori
- Source/Author:
- Gay G, Wagner DT, Keatinge-Clay AT, Gay DC.
pET-28 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-28 vector Sequence
LOCUS 40924_17791 5331 bp DNA circular SYN 18-DEC-2018
DEFINITION Expression vector pET-28, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5331)
AUTHORS Gay G, Wagner DT, Keatinge-Clay AT, Gay DC.
TITLE Rapid modification of the pET-28 expression vector for ligation
independent cloning using homologous recombination in Saccharomyces
cerevisiae
JOURNAL Plasmid 76, 66-71 (2014)
PUBMED 25304917
REFERENCE 2 (bases 1 to 5331)
AUTHORS Gay GR, Gay DC, Keatinge-Clay AT.
TITLE Direct Submission
JOURNAL Submitted (05-MAY-2014) Molecular Biosciences, University of Texas
at Austin, 1 University Station, Austin, TX 78712, USA
REFERENCE 3 (bases 1 to 5331)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 5331)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Plasmid 76,
66-71 (2014)"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(05-MAY-2014) Molecular Biosciences, University of Texas at Austin,
1 University Station, Austin, TX 78712, USA"
COMMENT SGRef: number: 3; type: "Journal Article"
COMMENT ##Assembly-Data-START##
Sequencing Technology :: Sanger dideoxy sequencing
##Assembly-Data-END##
FEATURES Location/Qualifiers
source 1..5331
/mol_type="other DNA"
/organism="synthetic DNA construct"
rep_origin 12..467
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
CDS complement(563..1375)
/codon_start=1
/label=KanR
/note="aminoglycoside phosphotransferase"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
rep_origin 1497..2085
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature complement(2271..2413)
/label=bom
/note="basis of mobility region from pBR322"
CDS complement(2518..2706)
/codon_start=1
/label=rop
/note="Rop protein, which maintains plasmids at low copy
number"
/translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
protein_bind complement(3481..3502)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(3518..4597)
/codon_start=1
/label=lacI
/note="lac repressor"
/translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
promoter complement(4598..4675)
/label=lacI promoter
promoter 4984..5002
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 5003..5027
/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 5042..5064
/label=RBS
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 5083..5100
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
CDS 5110..5127
/codon_start=1
/label=thrombin site
/note="thrombin recognition and cleavage site"
/translation="LVPRGS"
CDS 5175..5192
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
terminator 5259..5306
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
This page is informational only.