Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V003900 | pPV254 | 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
- Vector Name:
- pPV254
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6758 bp
- Type:
- PiggyBac_based_Ss_Act2_gfp_era1 vector
- Replication origin:
- ori
- Source/Author:
- Shao H, Li X, Nolan TJ, Massey HC Jr., Pearce EJ, Lok JB.
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 37℃
pPV254 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.
pPV254 vector Sequence
LOCUS 40924_35658 6758 bp DNA circular SYN 18-DEC-2018
DEFINITION PiggyBac_based_Ss_Act2_gfp_era1 vector pPV254, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6758)
AUTHORS Shao H, Li X, Nolan TJ, Massey HC Jr., Pearce EJ, Lok JB.
TITLE Transposon-mediated Chromosomal Integration of Transgenes in the
Parasitic Nematode Strongyloides ratti and Establishment of Stable
Transgenic Lines
JOURNAL PLoS Pathog. 8 (8), E1002871 (2012)
PUBMED 22912584
REFERENCE 2 (bases 1 to 6758)
AUTHORS Shao H, Li X, Nolan TJ, Massey HC Jr., Pearce EJ, Lok JB.
TITLE Direct Submission
JOURNAL Submitted (03-MAY-2012) Pathobiology, University of Pennsylvania,
3800 Spruce Street, Philadelphia, PA 19104, USA
REFERENCE 3 (bases 1 to 6758)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 6758)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "PLoS
Pathog."; date: "2012"; volume: "8"; issue: "8"; pages: "E1002871"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(03-MAY-2012) Pathobiology, University of Pennsylvania, 3800 Spruce
Street, Philadelphia, PA 19104, USA"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6758
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 24..128
/label=AmpR promoter
CDS 129..986
/label=AmpR
/note="beta-lactamase"
rep_origin 1160..1748
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
protein_bind 2036..2057
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 2072..2102
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2110..2126
/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)."
primer_bind 2134..2150
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
mobile_element 2641..2675
/mobile_element_type="transposon:piggyBac ITR"
/label=ITR
CDS complement(3315..3326)
/label=Factor Xa site
/note="Factor Xa recognition and cleavage site"
misc_recomb 3826..3830
/gene="act2"
/label=target sequence
/note="target sequence"
CDS join(4249..4419,4471..4624,4676..4839,4891..5118)
/codon_start=1
/product="GFP"
/label=GFP
/protein_id="AFN89784.1"
/translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
FICTTGKLPVPWPTLVTTFCYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
FVTAAGITHGMDELYK"
3'UTR 5138..5724
/gene="era1"
gene 5138..5724
/gene="era1"
/label=era1
/note="Ss-era1"
primer_bind 5738..5754
/label=SK primer
/note="common sequencing primer, one of multiple similar
variants"
mobile_element 5951..6013
/mobile_element_type="transposon:piggyBac ITR"
/label=ITR
misc_recomb 6014..6017
/label=target sequence
/note="target sequence"
primer_bind complement(6143..6159)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
rep_origin 6301..6756
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"