Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V010329 | pAbAi | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
To use pAbAi in a one-hybrid assay, clone one or more copies of a cis-acting element into the MCS. To facilitate recombination of the vector with the host’s ura3-52 locus, linearize the vector with either BbsI or BstBI, then introduce the linearized vector into competent yeast cells to integrate into the chromsome via homologous recombination between the vector’s URA3 gene and the ura3-52 locus of the yeast strain.
- Vector Name:
- pAbAi
- Antibiotic Resistance:
- Ampicillin
- Length:
- 4894 bp
- Type:
- Yeast Plasmids
- Replication origin:
- ori
- Host:
- Yeast
- Source/Author:
- Clontech
- Copy Number:
- High copy number
- Promoter:
- URA3
- Growth Strain(s):
- JM108
- Growth Temperature:
- 37℃
pAbAi 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.
pAbAi vector Sequence
LOCUS pAbAi. 4894 bp DNA circular SYN 01-JAN-1980
DEFINITION Yeast integrating vector for cloning a DNA bait sequence to be used
in a one-hybrid screen.
ACCESSION .
VERSION .
KEYWORDS pAbAi
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4894)
AUTHORS Clontech
TITLE Direct Submission
REFERENCE 2 (bases 1 to 4894)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
COMMENT Linearize with BstBI or BbsI to integrate at the URA3 locus.
FEATURES Location/Qualifiers
source 1..4894
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 1..45
/label=MCS
/note="MCS"
/note="multiple cloning site"
CDS 185..1387
/label=AurR
/note="aureobasidin A-resistant mutant of S. cerevisiae
inositol phosphorylceramide synthase (Aeed et al., 2009)"
primer_bind complement(1645..1661)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(1669..1685)
/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(1693..1723)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(1738..1759)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(2047..2635)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(2809..3666)
/label=AmpR
/note="beta-lactamase"
promoter complement(3667..3771)
/label=AmpR promoter
CDS complement(3878..4678)
/label=URA3
/note="orotidine-5'-phosphate decarboxylase, required for
uracil biosynthesis"
promoter complement(4679..4894)
/label=URA3 promoter