Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V016775 | Tet-pLKO.Puro_shGFP | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- Tet-pLKO.Puro_shGFP
- Antibiotic Resistance:
- Ampicillin
- Length:
- 8816 bp
- Type:
- RNA interference, Tetracycline inducible
- Replication origin:
- ori
- Host:
- Mammalian cells, Lentivirus
- Selection Marker:
- Puro
- Promoter:
- hPGK
- Growth Strain(s):
- Stbl3
- Growth Temperature:
- 37℃
Tet-pLKO.Puro_shGFP vector 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.
Tet-pLKO.Puro_shGFP vector Sequence
LOCUS V016775 8816 bp DNA circular SYN 01-JAN-1980
DEFINITION Exported.
ACCESSION V016775
VERSION V016775
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 8816)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..8816
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 1..227
/label="RSV promoter"
/note="Rous sarcoma virus enhancer/promoter"
LTR 228..408
/label="5' LTR (truncated)"
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 455..580
/label="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1073..1306
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 1491..1535
/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 1684..1725
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
promoter 1824..1918
/label="H1-2O2 promoter"
/note="doxycycline-inducible variant of the human H1 RNA
promoter (Henriksen et al., 2007)"
misc_feature 2019..2136
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 2185..2695
/label="hPGK promoter"
/note="human phosphoglycerate kinase 1 promoter"
intron 2704..3276
/label="beta-globin intron"
/note="intron from rabbit beta-globin gene"
promoter 3331..3349
/label="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 3360..3980
/label="TetR"
/note="tetracycline repressor TetR"
misc_feature 4015..4588
/label="IRES"
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
CDS 4608..5204
/label="PuroR"
/note="puromycin N-acetyltransferase"
LTR 5335..5568
/label="3' LTR (Delta-U3)"
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 5640..5774
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
rep_origin 5801..5936
/label="SV40 ori"
/note="SV40 origin of replication"
promoter complement(5957..5975)
/label="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(5985..6001)
/label="M13 fwd"
/note="common sequencing primer, one of multiple similar
variants"
rep_origin 6143..6598
/label="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 6624..6728
/label="AmpR promoter"
CDS 6729..7586
/label="AmpR"
/note="beta-lactamase"
rep_origin 7760..8348
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
protein_bind 8636..8657
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 8672..8702
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind 8710..8726
/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 8734..8750
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
promoter 8771..8789
/label="T3 promoter"
/note="promoter for bacteriophage T3 RNA polymerase"