LOCUS       Exported               11103 bp ds-DNA     circular SYN 20-DEC-2021
DEFINITION  2nd gen bicistronic lentiviral vector allows for simultaneous 
            expression of a transgene and EGFP marker to facilitate tracking of 
            transduced cells..
ACCESSION   .
VERSION     .
KEYWORDS    pWPI
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11103)
  TITLE     Trono Lab Constitutive Lentiviral Plasmids
REFERENCE   2  (bases 1 to 11103)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..11103
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     182..202
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     complement(272..289)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(443..1031)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(523..542)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1202..2062)
                     /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     1825..1844
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2063..2167)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2235..2253
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2291..2313)
                     /label=pGEX 3
                     /note="pGEX vectors, reverse primer"
     primer_bind     2413..2432
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2646..2665
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2955..2973
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2955..2972
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     LTR             2998..3178
                     /label=5 LTR (truncated)
                     /note="truncated 5 long terminal repeat (LTR) from HIV-1"
     protein_bind    complement(3198..3231)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    3375..3963
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3481..3492
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     3890..3910
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     primer_bind     3976..3992
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3977..3993
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(4072..4788)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(4072..4788)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4114..4135)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     4463..4482
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4722..4743
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     misc_feature    4791..5342
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4952..4971)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     5181..5198
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     misc_feature    5409..5526
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        complement(5574..6752)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          5583..6521
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(5606..5626)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             complement(7135..7179)
                     /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    7364..7597
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    8090..8215
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             8262..8895
                     /label=3 LTR
                     /note="3 long terminal repeat (LTR) from HIV-1"
     polyA_signal    9063..9197
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(9087..9106)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     9141..9160
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(9622..9642)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          9772..9837
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     promoter        complement(join(10971..11103,1..197))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(10985..11103,1..17)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(11039..11058)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"