LOCUS       Exported               11712 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Lentiviral expression of SARS-CoV-2 protein; transient expression 
            and generate lentivirus.
ACCESSION   .
VERSION     .
KEYWORDS    pLVX-EF1alpha-SARS-CoV-2-nsp12-2xStrep-IRES-Puro
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11712)
  AUTHORS   Gordon DE, et al.
  TITLE     A SARS-CoV-2-Human Protein-Protein Interaction Map Reveals Drug 
            Targets and Potential Drug Repurposing
  JOURNAL   BioRxiv March 2020
REFERENCE   2  (bases 1 to 11712)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BioRxiv 
            March 2020"
FEATURES             Location/Qualifiers
     source          1..11712
                     /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    2028..2143
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     promoter        2338..3519
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2568..3510
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3467..3487
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             6367..6390
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             6427..6450
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     misc_feature    6462..7035
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(6629..6646)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     6856..6875
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             7068..7667
                     /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(7068..7087)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7564..7584
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    7681..8269
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7734..7754)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8152..8163)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8476..9109
                     /label=3 LTR
                     /note="3 long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(9237..9253)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9237..9253)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(9250..9272)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    9261..9277
                     /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)."
     promoter        complement(9285..9315)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9330..9351
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(9468..9485)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(9639..10227)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9719..9738)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(10398..11258)
                     /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     11021..11040
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(11259..11363)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    11411..11545
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11448..11467)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11502..11521
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"