Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V000836 | p2CT-His-MBP-Lba_Cas13a_WT | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- p2CT-His-MBP-Lba_Cas13a_WT
- Antibiotic Resistance:
- Ampicillin
- Length:
- 10267 bp
- Type:
- Bacterial Expression
- Replication origin:
- ori
- Copy Number:
- High Copy
- Promoter:
- tet
- Cloning Method:
- Ligation Independent Cloning
p2CT-His-MBP-Lba_Cas13a_WT 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.
p2CT-His-MBP-Lba_Cas13a_WT vector Sequence
LOCUS V000836 10267 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V000836
VERSION V000836
KEYWORDS p2CT-His-MBP-Lba_Cas13a_WT
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 10267)
AUTHORS East-Seletsky A, O'Connell MR, Burstein D, Knott GJ, Doudna JA
TITLE RNA Targeting by Functionally Orthogonal Type VI-A CRISPR-Cas
Enzymes.
JOURNAL Mol Cell. 2017 May 4;66(3):373-383.e3. doi:
10.1016/j.molcel.2017.04.008.
PUBMED 28475872
REFERENCE 2 (bases 1 to 10267)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 10267)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1016/j.molcel.2017.04"; journalName: "Mol Cell"; date:
"2017-05-4- 4"; volume: "66"; issue: "3"; pages: "373-383.e3"
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..10267
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter complement(233..261)
/label="tet promoter"
/note="E. coli promoter for tetracycline efflux protein
gene"
primer_bind complement(288..306)
/label="pBRforEco"
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 374..478
/label="AmpR promoter"
CDS 479..1336
/label="AmpR"
/note="beta-lactamase"
rep_origin 1510..2098
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 2252..2269
/label="L4440"
/note="L4440 vector, forward primer"
misc_feature complement(2284..2423)
/label="bom"
/note="basis of mobility region from pBR322"
primer_bind 2509..2531
/label="pGEX 3'"
/note="pGEX vectors, reverse primer"
CDS complement(2528..2716)
/label="rop"
/note="Rop protein, which maintains plasmids at low copy
number"
primer_bind 4174..4193
/label="pBRrevBam"
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
promoter 4277..4295
/label="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
RBS 4384..4406
/label="RBS"
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 4426..4443
/label="6xHis"
/note="6xHis affinity tag"
CDS 4453..5553
/label="MBP"
/note="maltose binding protein from E. coli"
CDS 5608..5628
/label="TEV site"
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
CDS 5635..9945
/gene="cas13a"
/label="CRISPR-associated endoribonuclease Cas13a"
/note="CRISPR-associated endoribonuclease Cas13a from
Lachnospiraceae bacterium (strain NK4A179). Accession#:
P0DPB7"
terminator 10087..10134
/label="T7 terminator"
/note="transcription terminator for bacteriophage T7 RNA
polymerase"