Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V013996 | pGL3-TK-5UTR-BsmBI-Luciferase | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
pGL3 - TK - 5UTR - BsmBI - Luciferase is an important tool for gene expression regulation research. It uses luciferase activity to show the impact of 5′UTRs on gene expression and has wide applications.
Its vector construction includes a luciferase reporter gene, a TK promoter for driving transcription in various cell types, and BsmBI cloning sites for inserting 5′UTR fragments.
Functional advantages are studying gene expression regulation mechanisms of different genes’ 5′UTRs by detecting luciferase activity in cells, like in breast cancer - related genes research, and being suitable for high - throughput screening experiments to quickly identify significant 5′UTRs or regulatory elements for gene regulation network research.
- Vector Name:
- pGL3-TK-5UTR-BsmBI-Luciferase
- Antibiotic Resistance:
- Ampicillin
- Length:
- 5359 bp
- Type:
- Luciferase reporter
- Replication origin:
- ori
- Host:
- Mammalian cells
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 37℃
pGL3-TK-5UTR-BsmBI-Luciferase 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.
References
- Jewer M, Lee L, Leibovitch M, Zhang G, Liu J, Findlay SD, Vincent KM, Tandoc K, Dieters-Castator D, Quail DF, Dutta I, Coatham M, Xu Z, Puri A, Guan BJ, Hatzoglou M, Brumwell A, Uniacke J, Patsis C, Koromilas A, Schueler J, Siegers GM, Topisirovic I, Postovit LM. Translational control of breast cancer plasticity. Nat Commun. 2020 May 19;11(1):2498. doi: 10.1038/s41467-020-16352-z. PMID: 32427827; PMCID: PMC7237473.
pGL3-TK-5UTR-BsmBI-Luciferase vector Sequence
LOCUS 62056_11680 5359 bp DNA circular SYN 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5359)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..5359
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind 211..232
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 247..277
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 285..301
/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 309..325
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
promoter 346..364
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
promoter complement(412..430)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(440..456)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
CDS 629..2278
/codon_start=1
/label=luciferase
/note="firefly luciferase"
/translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
polyA_signal complement(2322..2443)
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin complement(2862..3450)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(3624..4481)
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(4482..4586)
/label=AmpR promoter
rep_origin 4613..5068
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
polyA_signal 5199..5247
/label=poly(A) signal
/note="synthetic polyadenylation signal"
misc_feature 5261..5352
/label=pause site
/note="RNA polymerase II transcriptional pause signal from
the human alpha-2 globin gene"