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pEBTet_NAD-Snifit-cyto vector (V016998)

Price Information

Cat No. Plasmid Name Availability Add to cart
V016998 pEBTet_NAD-Snifit-cyto 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:
pEBTet_NAD-Snifit-cyto
Antibiotic Resistance:
Ampicillin
Length:
13797 bp
Type:
Protein expression
Replication origin:
ori
Host:
Mammalian cells
Selection Marker:
Puro
Promoter:
RSV
Growth Strain(s):
DH5a
Growth Temperature:
37℃

pEBTet_NAD-Snifit-cyto vector Map

Copy Sequence View Map
pEBTet_NAD-Snifit-cyto13797 bp600120018002400300036004200480054006000660072007800840090009600102001080011400120001260013200Sepiapterin reductaseHaloTag(R)SNAP-tag(R)bGH poly(A) signalAmpR promoterAmpRoriSV40 promoterPuroRSV40 poly(A) signalRSV promoterbeta-globin intronT7 promoterTetRCMV enhancerCMV promotertet operator

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.

pEBTet_NAD-Snifit-cyto vector Sequence

LOCUS       V016998                13797 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V016998
VERSION     V016998
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 13797)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..13797
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             12..794
                     /gene="SPR"
                     /label="Sepiapterin reductase"
                     /note="Sepiapterin reductase from Homo sapiens. Accession#:
                     P35270"
     CDS             804..1691
                     /label="HaloTag(R)"
                     /note="modified bacterial dehalogenase that forms covalent
                     bonds with chloroalkane derivatives"
     CDS             1815..2360
                     /label="SNAP-tag(R)"
                     /note="human O6-alkylguanine-DNA-alkyltransferase variant
                     that forms covalent bonds with benzylguanine derivatives"
     polyA_signal    2428..2539
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     promoter        7619..7723
                     /label="AmpR promoter"
     CDS             7724..8581
                     /label="AmpR"
                     /note="beta-lactamase"
     rep_origin      8755..9343
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     promoter        9550..9847
                     /label="SV40 promoter"
                     /note="SV40 enhancer and early promoter"
     CDS             9924..10520
                     /label="PuroR"
                     /note="puromycin N-acetyltransferase"
     polyA_signal    10614..10748
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     promoter        11182..11443
                     /label="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     intron          11560..12132
                     /label="beta-globin intron"
                     /note="intron from rabbit beta-globin gene"
     promoter        12187..12205
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             12216..12836
                     /label="TetR"
                     /note="tetracycline repressor TetR"
     enhancer        13054..13433
                     /label="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        13434..13637
                     /label="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     protein_bind    13639..13657
                     /label="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"