LOCUS       Exported               11116 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Lentiviral vector for constitutive expression of the LaG17 GFP 
            nanobody synNotch Gal4DBDVP64 Receptor (PGK promoter).
ACCESSION   .
VERSION     .
KEYWORDS    pHR_pGK_LaG17_synNotch_Gal4VP64
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11116)
  AUTHORS   Morsut L, Roybal KT, Xiong X, Gordley RM, Coyle SM, Thomson M, Lim 
            WA
  TITLE     Engineering Customized Cell Sensing and Response Behaviors Using 
            Synthetic Notch Receptors.
  JOURNAL   Cell. 2016 Feb 11;164(4):780-91. doi: 10.1016/j.cell.2016.01.012. 
            Epub 2016 Jan 28.
  PUBMED    26830878
REFERENCE   2  (bases 1 to 11116)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cell.2016.01.012"; journalName: "Cell"; date: 
            "2016-02-11- 11"; volume: "164"; issue: "4"; pages: "780-91"
FEATURES             Location/Qualifiers
     source          1..11116
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     LTR             1..634
                     /label=3 LTR
                     /note="3 long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /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    2050..2167
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2227..2726
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2262..2285)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2659..2678
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      2774..2783
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2843..2872
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4712..4861
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     misc_feature    4980..5568
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5033..5053)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5451..5462)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5571..5587)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5572..5588)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5678..5911
                     /label=3 LTR (Delta-U3)
                     /note="self-inactivating 3 long terminal repeat (LTR) from
                     HIV-1"
     promoter        complement(5936..5954)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5937..5954)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(6244..6263)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(6477..6496)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6596..6618
                     /label=pGEX 3
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6656..6674)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6742..6846
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6847..7707
                     /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(7065..7084)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7878..8466
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8367..8386
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8620..8637
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(8707..8727)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        8712..9041
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      8892..9027
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     8954..8973
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     intron          10175..10240
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             10370..10390
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    10815..10949
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10852..10871)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10906..10925
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"