Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V003816 | pRDV | 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
The pRDV is a ribosome-display vector. pRDV contains all flanking DNA regions necessary for ribosome display: the T7-promoter, the ribosomal binding site and an in-frame tolA gene spacer. Hence, by simple ligation of the DNA encoding the combinatorial library into pRDV and by a PCR using this ligation mix as template, all features necessary for ribosome display are added to the library. The use of pRDV has the advantage that it always provides error-free library flanking regions and that it saves a number of working steps compared to the standard PCR approach for library generation.
- Vector Name:
- pRDV
- Antibiotic Resistance:
- Ampicillin
- Length:
- 4279 bp
- Type:
- Ribosome display vector
- Replication origin:
- ori
- Source/Author:
- Binz HK, Amstutz P, Kohl A, Stumpp MT, Briand C, Forrer P, Grutter MG, Pluckthun A.
pRDV vector 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
- Binz HK, Amstutz P, Kohl A, Stumpp MT, Briand C, Forrer P, Grütter MG, Plückthun A. High-affinity binders selected from designed ankyrin repeat protein libraries. Nat Biotechnol. 2004 May;22(5):575-82. doi: 10.1038/nbt962. Epub 2004 Apr 18. PMID: 15097997.
pRDV vector Sequence
LOCUS 40924_36443 4279 bp DNA circular SYN 18-DEC-2018
DEFINITION Ribosome display vector pRDV, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4279)
AUTHORS Binz HK, Amstutz P, Kohl A, Stumpp MT, Briand C, Forrer P, Grutter
MG, Pluckthun A.
TITLE High-affinity binders selected from designed ankyrin repeat protein
libraries
JOURNAL Nat. Biotechnol. 22 (5), 575-582 (2004)
PUBMED 15097997
REFERENCE 2 (bases 1 to 4279)
AUTHORS Amstutz P, Zahnd C, Binz KH, Stumpp MT, Forrer P, Pluckthun A.
TITLE Direct Submission
JOURNAL Submitted (20-JUN-2003) Biochemistry, University of Zurich,
Winterthurerstrasse 190, Zurich 8057, Switzerland
REFERENCE 3 (bases 1 to 4279)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 4279)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nat.
Biotechnol."; date: "2004"; volume: "22"; issue: "5"; pages:
"575-582"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(20-JUN-2003) Biochemistry, University of Zurich,
Winterthurerstrasse 190, Zurich 8057, Switzerland"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..4279
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 19..37
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
stem_loop 36..56
protein_bind 60..76
/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 87..109
/label=RBS
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 123..146
/label=FLAG
/note="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
CDS 156..941
/label=bla(M)
/note="beta-lactamase lacking the signal sequence"
misc_feature 975..1862
/label=tolA spacer
/note="tolA spacer"
terminator 1930..1977
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 2013..2468
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 2495..2599
/label=AmpR promoter
CDS 2600..3457
/label=AmpR
/note="beta-lactamase"
rep_origin 3631..4219
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"