Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V000401 | pEBTet_NAD-Snifit-mito | 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:
- pEBTet_NAD-Snifit-mito
- Antibiotic Resistance:
- Ampicillin
- Length:
- 14157 bp
- Type:
- Mammalian Expression
- Replication origin:
- ori
- Selection Marker:
- Puromycin
- Copy Number:
- High Copy
- Promoter:
- RSV
- Cloning Method:
- Gibson Cloning
- 5' Primer:
- CMV_Fwd
- 3' Primer:
- BGH_Rev
pEBTet_NAD-Snifit-mito 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.
pEBTet_NAD-Snifit-mito vector Sequence
LOCUS V000401 14157 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V000401
VERSION V000401
KEYWORDS pEBTet_NAD-Snifit-mito
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 14157)
AUTHORS Sallin O, Reymond L, Gondrand C, Raith F, Koch B, Johnsson K
TITLE Semisynthetic biosensors for mapping cellular concentrations of
nicotinamide adenine dinucleotides.
JOURNAL Elife. 2018 May 29;7. pii: 32638. doi: 10.7554/eLife.32638.
PUBMED 29809136
REFERENCE 2 (bases 1 to 14157)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 14157)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Elife.";
date: "2018-05-29"; pages: "
10.7554/eLife.32638"
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..14157
/mol_type="other DNA"
/organism="synthetic DNA construct"
CDS 12..86
/label="COX8 presequence"
/note="mitochondrial presequence of human cytochrome c
oxidase subunit VIII (Rizutto et al., 1989)"
CDS 99..170
/codon_start=1
/product="mitochondrial presequence of human cytochrome c
oxidase subunit VIII (Rizutto et al., 1989)"
/label="COX8 presequence"
/translation="SVLTPLLLRGLTGSARRLPVPRAK"
CDS 189..260
/codon_start=1
/product="mitochondrial presequence of human cytochrome c
oxidase subunit VIII (Rizutto et al., 1989)"
/label="COX8 presequence"
/translation="SVLTPLLLRGLTGSARRLPVPRAK"
CDS 273..344
/codon_start=1
/product="mitochondrial presequence of human cytochrome c
oxidase subunit VIII (Rizutto et al., 1989)"
/label="COX8 presequence"
/translation="SVLTPLLLRGLTGSARRLPVPRAK"
CDS 375..1154
/gene="SPR"
/label="Sepiapterin reductase"
/note="Sepiapterin reductase from Homo sapiens. Accession#:
P35270"
CDS 1164..2051
/label="HaloTag(R)"
/note="modified bacterial dehalogenase that forms covalent
bonds with chloroalkane derivatives"
CDS 2175..2720
/label="SNAP-tag(R)"
/note="human O6-alkylguanine-DNA-alkyltransferase variant
that forms covalent bonds with benzylguanine derivatives"
polyA_signal 2788..2899
/label="bGH poly(A) signal"
/note="bovine growth hormone polyadenylation signal"
rep_origin 3440..5229
/label="oriP"
/note="Epstein-Barr virus oriP replication origin (Yates et
al., 2000)"
primer_bind complement(7893..7911)
/label="pBRforEco"
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 7979..8083
/label="AmpR promoter"
CDS 8084..8941
/label="AmpR"
/note="beta-lactamase"
rep_origin 9115..9703
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(9892..9912)
/label="pBABE 3'"
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 10077..10210
/label="SV40 ori"
/note="SV40 origin of replication"
CDS 10284..10880
/label="PuroR"
/note="puromycin N-acetyltransferase"
polyA_signal 10974..11108
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
primer_bind complement(11246..11265)
/label="pRS-marker"
/note="pRS vectors, use to sequence yeast selectable
marker"
promoter 11542..11803
/label="RSV promoter"
/note="Rous sarcoma virus enhancer/promoter"
intron 11920..12492
/label="beta-globin intron"
/note="intron from rabbit beta-globin gene"
primer_bind 12500..12519
/label="pCAG-F"
/note="Rabbit beta-globin intron, for pCAG plasmids,
forward primer"
promoter 12547..12565
/label="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 12576..13196
/label="TetR"
/note="tetracycline repressor TetR"
polyA_signal 13238..13359
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
enhancer 13414..13793
/label="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
promoter 13794..13997
/label="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
protein_bind 13999..14017
/label="tet operator"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 14020..14038
/gene="tetO"
/label="tet operator"
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
primer_bind 14048..14072
/label="LNCX"
/note="Human CMV promoter, forward primer"