Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V010857 | pEarleyGate 205 | 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:
- pEarleyGate 205
- Antibiotic Resistance:
- Chloramphenicol
- Length:
- 12188 bp
- Type:
- Plant Vectors
- Replication origin:
- ori
- Source/Author:
- Earley KW, Haag JR, Pontes O, Opper K, Juehne T, Song K, Pikaard CS.
- Copy Number:
- High copy number
- Promoter:
- MAS
pEarleyGate 205 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.
pEarleyGate 205 vector Sequence
LOCUS pEarleyGate_205. 12188 bp DNA circular SYN 01-JAN-1980
DEFINITION Gateway(R)-compatible plant transformation vector with a C-terminal
TAP tag.
ACCESSION .
VERSION .
KEYWORDS pEarleyGate 205
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 12188)
AUTHORS Earley KW, Haag JR, Pontes O, Opper K, Juehne T, Song K, Pikaard CS.
TITLE Gateway-compatible vectors for plant functional genomics and
proteomics.
JOURNAL Plant J. 2006;45:616-29.
PUBMED 16441352
REFERENCE 2 (bases 1 to 12188)
AUTHORS Pikaard Lab
TITLE Direct Submission
REFERENCE 3 (bases 1 to 12188)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Plant J.";
date: "2006"; volume: "45"; pages: "616-29"
COMMENT SGRef: number: 2; type: "Journal Article"
COMMENT The sequence was corrected by inserting a G at position 8296.
FEATURES Location/Qualifiers
source 1..12188
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 1..25
/label=LB T-DNA repeat
/note="left border repeat from nopaline C58 T-DNA"
terminator complement(111..363)
/label=MAS terminator
/note="mannopine synthase terminator"
CDS complement(376..924)
/label=BlpR
/note="phosphinothricin acetyltransferase"
promoter complement(930..1310)
/label=MAS promoter
/note="mannopine synthase promoter (Velten et al., 1984)"
promoter 2306..2651
/label=CaMV 35S promoter
/note="strong constitutive promoter from cauliflower mosaic
virus"
protein_bind 2663..2782
/label=attR1
/note="recombination site for the Gateway(R) LR reaction"
promoter 2812..2842
/label=lac UV5 promoter
/note="E. coli lac promoter with an 'up' mutation"
CDS 2896..3552
/label=CmR
/note="chloramphenicol acetyltransferase"
CDS 3897..4199
/label=ccdB
/note="CcdB, a bacterial toxin that poisons DNA gyrase"
protein_bind complement(4243..4367)
/label=attR2
/note="recombination site for the Gateway(R) LR reaction"
CDS 4381..4458
/label=CBP
/note="calmodulin-binding peptide"
CDS 4486..4506
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
CDS 4546..4719
/label=ProtA
/note="IgG-binding unit of Staphylococcus aureus protein A"
CDS 4720..4893
/codon_start=1
/product="IgG-binding unit of Staphylococcus aureus protein
A"
/label=IgG-binding unit of Staphylococcus aureus prote
/note="ProtA"
/translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
AEAKKLNGAQAPK"
terminator 4959..5666
/label=OCS terminator
/note="octopine synthase terminator"
misc_feature 5910..5934
/label=RB T-DNA repeat
/note="right border repeat from nopaline C58 T-DNA"
CDS 7234..7860
/label=pVS1 StaA
/note="stability protein from the Pseudomonas plasmid pVS1
(Heeb et al., 2000)"
CDS 8292..9362
/label=pVS1 RepA
/note="replication protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
rep_origin 9431..9625
/label=pVS1 oriV
/note="origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
misc_feature 9969..10109
/label=bom
/note="bom"
/note="basis of mobility region from pBR322"
rep_origin complement(10295..10883)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(10973..11764)
/label=KanR
/note="aminoglycoside phosphotransferase"