Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001243 | pComb3XTT | 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
TG1 or XL1-blue cells are recommended for transformation.
- Vector Name:
- pComb3XTT
- Antibiotic Resistance:
- Ampicillin
- Length:
- 4766 bp
- Type:
- Bacterial Expression ; phage display
- Replication origin:
- ori
- Copy Number:
- High Copy
- Promoter:
- lacZ
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- AAG ACA GCT ATC GCG ATT GCA G
- 3' Primer:
- GCC CCC TTA TTA GCG TTT GCC ATC
- Growth Strain(s):
- TG1
- Growth Temperature:
- 37℃
pComb3XTT 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.
pComb3XTT vector Sequence
LOCUS 40924_12745 4766 bp DNA circular SYN 13-MAY-2021
DEFINITION Backbone of pComb3X containing TT Fab (light and heavy chains)
against tetanus toxin, used as phage display/expression control and
PCR template for human antibody constant regions (kappa and CH1).
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4766)
AUTHORS Andris-Widhopf J, Rader C, Steinberger P, Fuller R, Barbas CF 3rd
TITLE Methods for the generation of chicken monoclonal antibody fragments
by phage display.
JOURNAL J Immunol Methods. 2000 Aug 28;242(1-2):159-81.
PUBMED 10986398
REFERENCE 2 (bases 1 to 4766)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 4766)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "J Immunol
Methods."; date: "2000-08-28"; volume: "242(1-2)"; pages: "159-81"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..4766
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind 105..126
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 141..171
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 179..195
/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)."
sig_peptide 218..280
/label=OmpA signal peptide
/note="signal peptide from the E. coli outer membrane
protein OmpA"
CDS 602..919
/codon_start=1
/label=hIg-kappa-CL
/note="Human immunoglobulin kappa light chain constant
region"
/translation="TVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNA
LQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSLPVTKSFNRG
EC"
sig_peptide 953..1018
/label=pelB signal sequence
/note="leader peptide for secretion"
CDS 1733..1750
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
CDS 1757..1783
/codon_start=1
/label=HA
/note="HA (human influenza hemagglutinin) epitope tag"
/translation="YPYDVPDYA"
rep_origin complement(2377..2832)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 2859..2963
/label=AmpR promoter
CDS 2964..3821
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRR
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 3995..4583
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 4737..4754
/label=L4440
/note="L4440 vector, forward primer"