Tet-pLKO-puro vector (V012735)

Basic Vector Information

3rd generation lentiviral plasmid for inducible expression of shRNA; puromycin selection.

      • Vector Name:
      • Tet-pLKO-puro
      • Antibiotic Resistance:
      • Ampicillin
      • Length:
      • 10633 bp
      • Type:
      • RNAi
      • Source/Author:
      • Dmitri Wiederschain
      • Selection Marker:
      • Puromycin
      • Copy Number:
      • High copy number
      • Expression Method:
      • Constiutive, Stable / Transient

Tet-pLKO-puro vector Vector Map

Tet-pLKO-puro10633 bp5001000150020002500300035004000450050005500600065007000750080008500900095001000010500beta-globin intronT7 promoterTetRIRESPuroR3' LTR (Delta-U3)SV40 poly(A) signalSV40 oriT7 promoterM13 fwdf1 oriAmpR promoterAmpRoriCAP binding sitelac promoterlac operatorM13 revT3 promoterRSV promoter5' LTR (truncated)HIV-1 PsiRREtet operatortet operatorcPPT/CTS

Plasmid Resuspension 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 ℃.

Tet-pLKO-puro vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       Exported               10633 bp ds-DNA     circular SYN 11-SEP-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    Tet-pLKO-puro
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10633)
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..10633
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     intron          516..1088
                     /note="beta-globin intron"
                     /note="intron from rabbit beta-globin gene"
     promoter        1143..1161
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1172..1789
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /note="TetR"
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESG"
     misc_feature    1827..2400
                     /note="IRES"
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     CDS             2420..3019
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /note="PuroR"
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     LTR             3147..3380
                     /note="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3452..3573
                     /note="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      3613..3748
                     /note="SV40 ori"
                     /note="SV40 origin of replication"
     promoter        complement(3769..3787)
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3797..3813)
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      3955..4410
                     /direction=RIGHT
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4436..4540
                     /gene="bla"
                     /note="AmpR promoter"
     CDS             4541..5401
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5572..6160
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    6448..6469
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding site"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6484..6514
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    6522..6538
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="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     6546..6562
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        6583..6601
                     /note="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6629..6855
                     /note="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6856..7036
                     /note="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7083..7208
                     /note="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7701..7934
                     /note="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     protein_bind    8500..8518
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8528..8546
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    10464..10581
                     /note="cPPT/CTS"
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        10630..507
                     /note="hPGK promoter"
                     /note="human phosphoglycerate kinase 1 promoter"

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