LOCUS       Exported                9637 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  3rd gen lentiviral Luciferase expression vector, CMV promoter, Puro.
ACCESSION   .
VERSION     .
KEYWORDS    pLenti CMV Puro LUC (w168-1)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9637)
  AUTHORS   Campeau E, Ruhl VE, Rodier F, Smith CL, Rahmberg BL, Fuss JO, 
            Campisi J, Yaswen P, Cooper PK, Kaufman PD.
  TITLE     A versatile viral system for expression and depletion of proteins in
            mammalian cells.
  JOURNAL   PLoS One. 2009 Aug 6;4(8):e6529.
  PUBMED    19657394
REFERENCE   2  (bases 1 to 9637)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS One.";
            date: "2009-08-6"; volume: "4(8)"; pages: "e6529"
FEATURES             Location/Qualifiers
     source          1..9637
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     505..529
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    613..637
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             643..2295
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(709..727)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     protein_bind    complement(2302..2326)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2357..2945
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2410..2430)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2828..2839)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        2959..3458
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2994..3017)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3391..3410
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3479..4078
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3479..3498)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3975..3995
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4216..4449
                     /label=3 LTR (Delta-U3)
                     /note="self-inactivating 3 long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4521..4642
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4558..4577)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4612..4631
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4682..4817
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4744..4763
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(4837..4856)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4838..4856)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4866..4883)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4866..4882)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4875..4897)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5024..5479
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5111..5130)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5321..5342
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5505..5609
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5610..6470
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5828..5847)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6641..7229
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7130..7149
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7383..7400
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7517..7538
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        7553..7583
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7591..7607
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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     7596..7618
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7615..7631
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7615..7631
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7650..7670
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7652..7670
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        7698..7924
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             7925..8105
                     /label=5 LTR (truncated)
                     /note="truncated 5 long terminal repeat (LTR) from HIV-1"
     misc_feature    8152..8277
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    8770..9003
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             9188..9232
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    9499..9615
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"