Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V012813 | pAIDA1 | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
A construct with the AIDA surface expression system for display of a passenger protein flanked by His and Myc tags
- Vector Name:
- pAIDA1
- Antibiotic Resistance:
- Chloramphenicol
- Length:
- 5561 bp
- Type:
- E.coli expression plasmid
- Replication origin:
- p15A ori
- Source/Author:
- Gen Larsson
- Copy Number:
- Low copy number
- Promoter:
- lacUV5
- Cloning Method:
- Enzyme Cut
- 5' Primer:
- GAGCGGATAACAATTTCACACAGG
- Growth Strain(s):
- DH5alpha
- Growth Temperature:
- 37℃
pAIDA1 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.
References
- Jarmander J, Gustavsson M, Do TH, Samuelson P, Larsson G. A dual tag system for facilitated detection of surface expressed proteins in Escherichia coli. Microb Cell Fact. 2012 Sep 3;11:118.
pAIDA1 vector Sequence
LOCUS pAIDA1. 5561 bp DNA circular SYN 11-MAY-2023
DEFINITION A construct with the AIDA surface expression system for display of a
passenger protein flanked by His and Myc tags.
ACCESSION .
VERSION .
KEYWORDS pAIDA1
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5561)
AUTHORS Jarmander J, Gustavsson M, Do TH, Samuelson P, Larsson G
TITLE A dual tag system for facilitated detection of surface expressed
proteins in Escherichia coli.
JOURNAL Microb Cell Fact. 2012 Sep 3;11:118. doi: 10.1186/1475-2859-11-118.
PUBMED 22943700
REFERENCE 2 (bases 1 to 5561)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Microb Cell
Fact."; date: "2012-09-3"; volume: "11:118. doi"; pages: "
10.1186/1475-2859-11-118"
FEATURES Location/Qualifiers
source 1..5561
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 1..31
/label=lac UV5 promoter
/note="E. coli lac promoter with an 'up' mutation"
protein_bind 39..55
/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)."
CDS 77..229
/codon_start=1
/label=SP
/note="SP"
/translation="MNKAYSIIWSHSRQAWIVASELARGHGFVLAKNTLLVLAVVSTIG
NAFAVD"
CDS 230..247
/label=6xHis
/note="6xHis affinity tag"
CDS 248..271
/codon_start=1
/label=HRV3C
/note="HRV3C"
/translation="LEALFQGP"
CDS 299..319
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
CDS 320..349
/label=Myc
/note="Myc (human c-Myc proto-oncogene) epitope tag"
CDS 356..1849
/codon_start=1
/label=ADIA
/note="ADIA"
/translation="VNNNGSIVINNSIINGNITNDADLSFGTAKLLSATVNGSLVNNKN
IILNPTKESAAAIGNTLTVSNYTGTPGSVISLGGVLEGDNSLTDRLVVKGNTSGQSDIV
YVNEDGSGGQTRDGINIISVEGNSDAEFSLKNRVVAGAYDYTLQKGNESGTDNKGWYLT
SHLPTSDTRQYRPENGSYATNMALANSLFLMDLNERKQFRAMSDNTQPESASVWMKITG
GISSGKLNDGQNKTTTNQFINQLGGDIYKFHAEQLGDFTLGIMGGYANAKGKTINYTSN
KAARNTLDGYSVGVYGTWYQNGENATGLFAETWMQYNWFNASVKGDGLEEEKYNLNGLT
ASAGGGYNLNVHTWTSPEGITGEFWLQPHLQAVWMGVTPDTHQEDNGTVVQGAGKNNIQ
TKAGIRASWKVKSTLDKDTGRRFRPYIEANWIHNTHEFGVKMSDDSQLLSGSRNQGEIK
TGIEGVITQNLSVNGGVAYQAGGHGSNAISGALGIKYSF"
rep_origin 2344..2889
/label=p15A ori
/note="Plasmids containing the medium-copy-number p15A
origin of replication can be propagated in E. coli cells
that contain a second plasmid with the ColE1 origin."
primer_bind 3006..3023
/label=L4440
/note="L4440 vector, forward primer"
primer_bind complement(3473..3492)
/label=pENTR-R
/note="pENTR vectors, reverse primer"
CDS 3659..4315
/label=CmR
/note="chloramphenicol acetyltransferase"
promoter 4331..4408
/label=lacI promoter
CDS 4409..5488
/label=lacI
/note="lac repressor"