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pCAG HRP-TM vector (V006519)

Price Information

Cat No. Plasmid Name Availability Add to cart
V006519 pCAG HRP-TM In stock, 1 week for quality controls

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

Basic Vector Information

Vector Name:
pCAG HRP-TM
Antibiotic Resistance:
Ampicillin
Length:
5469 bp
Type:
Mammalian Expression
Replication origin:
ori
Promoter:
CAG
Cloning Method:
Restriction Enzyme
5' Primer:
pCAG-F

pCAG HRP-TM vector Map

Copy Sequence View Map
pCAG HRP-TM5469 bp60012001800240030003600420048005400CMV enhancerchimeric intronpCAG-FIg-kappa leaderHAMycPDGFR-beta TM domainM13 revlac operatorlac promoterCAP binding siteSV40 promoterSV40 poly(A) signalL4440oriAmpRAmpR promoter

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.

pCAG HRP-TM vector Sequence

LOCUS       40924_8621        5469 bp DNA     circular SYN 09-JUL-2021
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5469)
  AUTHORS   Rhee HW, Zou P, Udeshi ND, Martell JD, Mootha VK, Carr SA, Ting AY
  TITLE     Proteomic Mapping of Mitochondria in Living Cells via Spatially 
            Restricted Enzymatic Tagging.
  JOURNAL   Science. 2013 Jan 31.
  PUBMED    23371551
REFERENCE   2  (bases 1 to 5469)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 5469)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science.
            2013 Jan 31."
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5469
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          614..1630
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and
                     rabbit beta-globin"
     primer_bind     1638..1657
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids,
                     forward primer"
     sig_peptide     1729..1791
                     /label=Ig-kappa leader
                     /note="leader sequence from mouse immunoglobulin kappa 
                     light
                     chain"
     CDS             1792..1818
                     /codon_start=1
                     /label=HA
                     /note="HA (human influenza hemagglutinin) epitope tag"
                     /translation="YPYDVPDYA"
     CDS             2770..2799
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2803..2949
                     /codon_start=1
                     /label=PDGFR-beta TM domain
                     /note="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(2971..2987)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2995..3011)
                     /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)."
     promoter        complement(3019..3049)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3064..3085)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        3143..3339
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     polyA_signal    3345..3479
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3547..3564)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3718..4306)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4480..5337)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5338..5442)
                     /label=AmpR promoter