Tet-pLKO-Hygro vector (V013552)

Price Information

Cat No. Plasmid Name Availability Add to cart
V013552 Tet-pLKO-Hygro 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:
Tet-pLKO-Hygro
Antibiotic Resistance:
Ampicillin
Length:
11003 bp
Type:
RNA interference, Tetracycline inducible
Replication origin:
ori
Host:
Mammalian cells, Lentivirus
Selection Marker:
Hyg
Promoter:
hPGK
Growth Strain(s):
Stbl3
Growth Temperature:
37℃

Tet-pLKO-Hygro vector Vector Map

Tet-pLKO-Hygro11003 bp500100015002000250030003500400045005000550060006500700075008000850090009500100001050011000hPGK promoterbeta-globin intronT7 promoterTetRIRESHygR3' LTR (Delta-U3)SV40 poly(A) signalSV40 oriT7 promoterf1 oriAmpR promoterAmpRoriCAP binding sitelac promoterT3 promoterRSV promoter5' LTR (truncated)HIV-1 PsiRREgp41 peptideProtein TatH1-2O2 promoterhPGK promotercPPT/CTS

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.

Tet-pLKO-Hygro vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       V013552                11003 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V013552
VERSION     V013552
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 11003)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..11003
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        7..507
                     /label="hPGK promoter"
                     /note="human phosphoglycerate kinase 1 promoter"
     intron          516..1088
                     /label="beta-globin intron"
                     /note="intron from rabbit beta-globin gene"
     promoter        1143..1161
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1172..1792
                     /label="TetR"
                     /note="tetracycline repressor TetR"
     misc_feature    1827..2400
                     /label="IRES"
                     /note="internal ribosome entry site (IRES) of the
                     encephalomyocarditis virus (EMCV)"
     CDS             2420..3442
                     /label="HygR"
                     /note="aminoglycoside phosphotransferase from E. coli"
     LTR             3517..3750
                     /label="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3822..3956
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      3983..4118
                     /label="SV40 ori"
                     /note="SV40 origin of replication"
     promoter        complement(4139..4157)
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      4325..4780
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        4806..4910
                     /label="AmpR promoter"
     CDS             4911..5768
                     /label="AmpR"
                     /note="beta-lactamase"
     rep_origin      5942..6530
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     protein_bind    6818..6839
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        6854..6884
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     promoter        6953..6971
                     /label="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6999..7225
                     /label="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             7226..7406
                     /label="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7453..7578
                     /label="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    8071..8304
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             8489..8533
                     /label="gp41 peptide"
                     /note="antigenic peptide corresponding to amino acids 655
                     to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
                     al., 2013)"
     CDS             8682..8723
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
                     /label="Protein Tat"
     promoter        8822..8916
                     /label="H1-2O2 promoter"
                     /note="doxycycline-inducible variant of the human H1 RNA
                     promoter (Henriksen et al., 2007)"
     promoter        9999..10507
                     /label="hPGK promoter"
                     /note="human phosphoglycerate kinase 1 promoter"
     misc_feature    10834..10951
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"