Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V010577 | pNIC28-Bsa4 | In stock, instant shipping |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pNIC28-Bsa4
- Antibiotic Resistance:
- Kanamycin
- Length:
- 7284 bp
- Type:
- Structural Genomics Vectors
- Replication origin:
- ori
- Source/Author:
- Savitsky P, Bray J, Cooper CD, Marsden BD, Mahajan P, Burgess-Brown
- Copy Number:
- High copy number
- Promoter:
- sacB
pNIC28-Bsa4 vector Map
Plasmid 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 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
pNIC28-Bsa4 vector Sequence
LOCUS 40924_33252 7284 bp DNA circular SYN 18-DEC-2018
DEFINITION Expression vector pNIC28-Bsa4, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7284)
AUTHORS Stols L, Gu M, Dieckman L, Raffen R, Collart FR, Donnelly MI.
TITLE A new vector for high-throughput, ligation-independent cloning
encoding a tobacco etch virus protease cleavage site
JOURNAL Protein Expr. Purif. 25 (1), 8-15 (2002)
PUBMED 12071693
REFERENCE 2 (bases 1 to 7284)
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 (2011) In press
PUBMED 22027370
REFERENCE 3 (bases 1 to 7284)
AUTHORS Gileadi O, Burgess-Brown N, Loppnau P.
TITLE Direct Submission
JOURNAL Submitted (25-DEC-2006) Structural Genomics Consortium, University
of Oxford, Botnar Research Centre, Oxford OX3 7LD, UK
REFERENCE 4 (bases 1 to 7284)
TITLE Direct Submission
REFERENCE 5 (bases 1 to 7284)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Protein
Expr. Purif."; date: "2002"; volume: "25"; issue: "1"; pages: "8-15"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "N
Biotechnol (2011) In press"
COMMENT SGRef: number: 3; type: "Journal Article"; journalName: "Submitted
(25-DEC-2006) Structural Genomics Consortium, University of Oxford,
Botnar Research Centre, Oxford OX3 7LD, UK"
COMMENT SGRef: number: 4; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..7284
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 1..19
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 20..44
/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 59..81
/label=RBS
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 92..109
/label=6xHis
/note="6xHis affinity tag"
CDS 134..154
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
promoter 170..615
/label=sacB promoter
/note="sacB promoter and control region"
CDS 616..2034
/label=SacB
/note="secreted levansucrase that renders bacterial growth
sensitive to sucrose"
misc_feature complement(2083..2097)
/note="3'-LIC sequence; add complement to downstream PCR
primer"
CDS 2145..2162
/label=6xHis
/note="6xHis affinity tag"
terminator 2229..2276
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 2313..2768
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
CDS complement(2864..3676)
/label=KanR
/note="aminoglycoside phosphotransferase"
rep_origin 3798..4386
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature complement(4572..4714)
/label=bom
/note="basis of mobility region from pBR322"
CDS complement(4819..5007)
/label=rop
/note="Rop protein, which maintains plasmids at low copy
number"
protein_bind complement(5782..5803)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(5819..6898)
/label=lacI
/note="lac repressor"
promoter complement(6899..6976)
/label=lacI promoter