Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V000222 | pBbA5c-MevT(CO)-T1-MBIS(CO, ispA) | 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:
- pBbA5c-MevT(CO)-T1-MBIS(CO, ispA)
- Antibiotic Resistance:
- Chloramphenicol
- Length:
- 13541 bp
- Type:
- Bacterial Expression, Synthetic Biology
- Replication origin:
- p15A ori
- Copy Number:
- Low Copy
- Promoter:
- LacUV5, pTrc
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- gatcgtttaggcacccca
- 3' Primer:
- ATTAATAAGATGATC
pBbA5c-MevT(CO)-T1-MBIS(CO, ispA) 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.
pBbA5c-MevT(CO)-T1-MBIS(CO, ispA) vector Sequence
LOCUS V000222 13541 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V000222
VERSION V000222
KEYWORDS pBbA5c-MevT(CO)-T1-MBIS(CO, ispA)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 13541)
AUTHORS Peralta-Yahya PP, Ouellet M, Chan R, Mukhopadhyay A, Keasling JD,
Lee TS
TITLE Identification and microbial production of a terpene-based advanced
biofuel.
JOURNAL Nat Commun. 2011 Sep 27;2:483. doi: 10.1038/ncomms1494.
PUBMED 21952217
REFERENCE 2 (bases 1 to 13541)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 13541)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nat
Commun."; date: "2011-09-27"; pages: "
10.1038/ncomms1494"
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..13541
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind complement(15..36)
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(52..1131)
/label="lacI"
/note="lac repressor"
promoter complement(1132..1209)
/label="lacI promoter"
primer_bind 1415..1434
/label="pBRrevBam"
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
promoter 1520..1550
/label="lac UV5 promoter"
/note="E. coli lac promoter with an 'up' mutation"
protein_bind 1558..1574
/label="lac operator"
/bound_moiety="lac repressor encoded by lacI"
/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)."
CDS 1614..2795
/gene="atoB"
/label="Acetyl-CoA acetyltransferase"
/note="Acetyl-CoA acetyltransferase from Escherichia coli
(strain K12). Accession#: P76461"
CDS 2816..4288
/gene="ERG13"
/label="Hydroxymethylglutaryl-CoA synthase"
/note="Hydroxymethylglutaryl-CoA synthase from
Saccharomyces cerevisiae (strain ATCC 204508 / S288c).
Accession#: P54839"
CDS 5220..5231
/label="Factor Xa site"
/note="Factor Xa recognition and cleavage site"
terminator 5849..5920
/label="rrnB T1 terminator"
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 5936..5963
/label="T7Te terminator"
/note="phage T7 early transcription terminator"
promoter 6175..6204
/label="trc promoter"
/note="strong E. coli promoter; hybrid between the trp and
lac UV5 promoters"
protein_bind 6212..6228
/label="lac operator"
/bound_moiety="lac repressor encoded by lacI"
/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)."
CDS 6254..7582
/gene="ERG12"
/label="Mevalonate kinase"
/note="Mevalonate kinase from Saccharomyces cerevisiae
(strain ATCC 204508 / S288c). Accession#: P07277"
CDS 7606..8958
/gene="ERG8"
/label="Phosphomevalonate kinase"
/note="Phosphomevalonate kinase from Saccharomyces
cerevisiae (strain ATCC 204508 / S288c). Accession#:
P24521"
CDS 8982..10169
/gene="MVD1"
/label="Diphosphomevalonate decarboxylase"
/note="Diphosphomevalonate decarboxylase from Saccharomyces
cerevisiae (strain ATCC 204508 / S288c). Accession#:
P32377"
CDS 10193..10738
/gene="idi"
/label="Isopentenyl-diphosphate Delta-isomerase"
/note="Isopentenyl-diphosphate Delta-isomerase from
Escherichia coli (strain K12 / DH10B). Accession#: B1XEH6"
CDS 10762..11658
/gene="ispA"
/label="Farnesyl diphosphate synthase"
/note="Farnesyl diphosphate synthase from Escherichia coli
(strain K12). Accession#: P22939"
terminator 11706..11777
/label="rrnB T1 terminator"
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 11793..11820
/label="T7Te terminator"
/note="phage T7 early transcription terminator"
primer_bind complement(11848..11865)
/label="L4440"
/note="L4440 vector, forward primer"
rep_origin complement(11982..12527)
/direction=LEFT
/label="p15A ori"
/note="Plasmids containing the medium-copy-number p15A
origin of replication can be propagated in E. coli cells
that contain a second plasmid with the ColE1 origin."
terminator complement(12641..12735)
/label="lambda t0 terminator"
/note="transcription terminator from phage lambda"
CDS complement(12759..13415)
/label="CmR"
/note="chloramphenicol acetyltransferase"
promoter complement(13416..13518)
/label="cat promoter"
/note="promoter of the E. coli cat gene encoding
chloramphenicol acetyltransferase"