pGL3-TK-5UTR-BsmBI-Luciferase vector (V013996)

COA reports Sequencing Results MSDS Download GenBank File(.gb)

Price Information

Cat No.	Plasmid Name	Availability	Add to cart
V013996	pGL3-TK-5UTR-BsmBI-Luciferase	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

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

Copy Sequence View 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"