pGL4.44[luc2P/AP1 RE/Hygro] vector (V011506) Gene synthesis in pGL4.44[luc2P/AP1 RE/Hygro] backbone

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Price Information

Cat No.	Plasmid Name	Availability	Buy one, get one free! (?)
V011506	pGL4.44[luc2P/AP1 RE/Hygro]	In stock, instant shipping

Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

pGL4.44 [luc2P/AP1RE/Hygro] was employed to assess the impact of HBx protein on AP1 transcription factor activity, quantifying MAPK/JNK pathway activation via luciferase expression levels.

Vector Name:: pGL4.44[luc2P/AP1 RE/Hygro]

Antibiotic Resistance:: Ampicillin

Length:: 6052 bp

Type:: Luciferase Vectors

Replication origin:: ori

Source/Author:: Promega

Copy Number:: High copy number

Promoter:: minP

Growth Strain(s):: Top10

pGL4.44[luc2P/AP1 RE/Hygro] 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

Hayashi M, Deng L, Chen M, Gan X, Shinozaki K, Shoji I, Hotta H. Interaction of the hepatitis B virus X protein with the lysine methyltransferase SET and MYND domain-containing 3 induces activator protein 1 activation. Microbiol Immunol. 2016 Jan;60(1):17-25. doi: 10.1111/1348-0421.12345. PMID: 26616333.

pGL4.44[luc2P/AP1 RE/Hygro] vector Sequence

LOCUS       pGL4.44[luc2P_AP        6052 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Vector with a minimal promoter and an AP-1 response element for 
            studying cell signaling using destabilized luciferase.
ACCESSION   .
VERSION     .
KEYWORDS    pGL4.44[luc2P AP1 RE Hygro]
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6052)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 6052)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6052
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    105..153
                     /label=poly(A) signal
                     /note="synthetic polyadenylation signal"
     misc_feature    167..258
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from
                     the human alpha-2 globin gene"
     protein_bind    285..332
                     /label=activator protein 1 (AP-1) binding site
                     /bound_moiety="activator protein 1 (AP-1)"
                     /note="AP-1 RE"
                     /note="AP-1 response element (6 copies)"
     promoter        377..408
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     CDS             441..2090
                     /label=luciferase
                     /note="firefly luciferase"
     CDS             2094..2213
                     /label=hPEST
                     /note="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
     polyA_signal    complement(2265..2386)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        2580..2937
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2968..4002
                     /label=HygR
                     /note="hygromycin B phosphotransferase"
     polyA_signal    4029..4077
                     /label=synthetic polyadenylation signal
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(4404..4992)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5195..6052)
                     /label=AmpR
                     /note="beta-lactamase"