Basic Vector Information
- Vector Name:
- pNIC-Bio3 plasmid pNIC-Bio3
- Antibiotic Resistance:
- Kanamycin
- Length:
- 7335 bp
- Type:
- Expression vector
- Replication origin:
- ori
- Source/Author:
- Keates T, Cooper CD, Savitsky P, Allerston CK, Phillips C, Hammarstrom M, Daga N, Berridge G, Mahajan P, Burgess-Brown NA, Muller S, Graslund S, Gileadi O.
- Promoter:
- sacB
pNIC-Bio3 plasmid pNIC-Bio3 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 ℃.
pNIC-Bio3 plasmid pNIC-Bio3 vector Sequence
LOCUS 40924_33212 7335 bp DNA circular SYN 18-DEC-2018
DEFINITION Expression vector pNIC-Bio3 plasmid pNIC-Bio3, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7335)
AUTHORS Keates T, Cooper CD, Savitsky P, Allerston CK, Phillips C,
Hammarstrom M, Daga N, Berridge G, Mahajan P, Burgess-Brown NA,
Muller S, Graslund S, Gileadi O.
TITLE Expressing the human proteome for affinity proteomics: optimising
expression of soluble protein domains and in vivo biotinylation
JOURNAL N Biotechnol 29 (5), 515-525 (2012)
PUBMED 22027370
REFERENCE 2 (bases 1 to 7335)
AUTHORS Burgess-Brown NA, Gileadi O.
TITLE Direct Submission
JOURNAL Submitted (27-SEP-2011) Structural Genomics Consortium, SGC,
University of Oxford, ORCRB, Roosevelt Drive, Oxford OX3 7DQ, UK
REFERENCE 3 (bases 1 to 7335)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 7335)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "N
Biotechnol"; date: "2012"; volume: "29"; issue: "5"; pages:
"515-525"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(27-SEP-2011) Structural Genomics Consortium, SGC, University of
Oxford, ORCRB, Roosevelt Drive, Oxford OX3 7DQ, UK"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..7335
/mol_type="other DNA"
/organism="synthetic DNA construct"
RBS 12..34
/label=RBS
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 45..62
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
CDS 87..107
/codon_start=1
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
/translation="ENLYFQS"
promoter 123..568
/label=sacB promoter
/note="sacB promoter and control region"
CDS 569..1987
/codon_start=1
/label=SacB
/note="secreted levansucrase that renders bacterial growth
sensitive to sucrose"
/translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
VKDSILEQGQLTVNK"
CDS 2052..2096
/codon_start=1
/label=AviTag(TM)
/note="peptide tag that allows for enzymatic biotinylation"
/translation="GLNDIFEAQKIEWHE"
CDS 2149..2166
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
terminator 2233..2280
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 2317..2772
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
CDS complement(2868..3680)
/codon_start=1
/label=KanR
/note="aminoglycoside phosphotransferase"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
rep_origin 3802..4390
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature complement(4576..4718)
/label=bom
/note="basis of mobility region from pBR322"
CDS complement(4823..5011)
/codon_start=1
/label=rop
/note="Rop protein, which maintains plasmids at low copy
number"
/translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
protein_bind complement(5786..5807)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(5823..6902)
/codon_start=1
/label=lacI
/note="lac repressor"
/translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
promoter complement(6903..6980)
/label=lacI promoter
promoter 7289..7307
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 7308..7332
/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)."
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