LOCUS       sequence-366581-        8655 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Doxycycline-dependent expression of NS3/4A protease gene in 
            mammalian cells by retroviral transduction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8655)
  AUTHORS   Mosallanejad K, Kennedy SN, Bahleda KM, Slavik KM, Zhou W, Govande 
            AA, Hancks DC, Kranzusch PJ, Kagan JC
  TITLE     Species-specific self-DNA detection mechanisms by mammalian cyclic 
            GMP-AMP synthases.
  JOURNAL   Sci Immunol. 2023 Jan 20;8(79):eabp9765. doi: 
            10.1126/sciimmunol.abp9765. Epub 2023 Jan 20.
  PUBMED    36662885
REFERENCE   2  (bases 1 to 8655)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci 
            Immunol."; date: "2023-01-20"; volume: "8(79):eabp9765. doi"; pages:
            " 10.1126/sciimmunol.abp9765. Epub 2023 Jan 20"
COMMENT     Sequence Label: pTRE3G-NS3/4A
FEATURES             Location/Qualifiers
     source          1..8655
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        310..513
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             514..689
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    752..951
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1152..1568
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1498..1520
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1536..1552
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        1608..1957
                     /label=TRE3GV promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, optimized for 
                     retroviral and lentiviral vectors"
     protein_bind    1616..1634
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1652..1670
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1688..1706
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1724..1742
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1760..1778
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1796..1814
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1832..1850
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1896..1920
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1964..1973
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        4061..4560
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4096..4119)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     4493..4512
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             4581..5180
                     /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(4581..4600)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5077..5097
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             5361..5786
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(5889..5911)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6047..6067)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6052..6381
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6232..6367
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6403..6422
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6503..6520)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6674..7262)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6754..6773)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7433..8224,8225..8293))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8056..8075
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8294..8398)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8466..8484
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(8584..8655,1..309)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
       61 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      121 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      181 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgctctg gcattatgcc
      241 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      301 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      361 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      421 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      481 cgtgtacggt gggaggtcta tataagcaga gctcaataaa agagcccaca acccctcact
      541 cggcgcgcca gtcttccgat agactgcgtc gcccgggtac ccgtattccc aataaagcct
      601 cttgctgttt gcatccgaat cgtggtctcg ctgttccttg ggagggtctc ctctgagtga
      661 ttgactaccc acgacggggg tctttcattt gggggctcgt ccgggatttg gagacccctg
      721 cccagggacc accgacccac caccgggagg taagctggcc agcaacttat ctgtgtctgt
      781 ccgattgtct agtgtctatg tttgatgtta tgcgcctgcg tctgtactag ttagctaact
      841 agctctgtat ctggcggacc cgtggtggaa ctgacgagtt ctgaacaccc ggccgcaacc
      901 ctgggagacg tcccagggac tttgggggcc gtttttgtgg cccgacctga ggaagggagt
      961 cgatgtggaa tccgaccccg tcaggatatg tggttctggt aggagacgag aacctaaaac
     1021 agttcccgcc tccgtctgaa tttttgcttt cggtttggaa ccgaagccgc gcgtcttgtc
     1081 tgctgcagcg ctgcagcatc gttctgtgtt gtctctgtct gactgtgttt ctgtatttgt
     1141 ctgaaaatta gggccagact gttaccactc ccttaagttt gaccttaggt cactggaaag
     1201 atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa gaagagacgt tgggttacct
     1261 tctgctctgc agaatggcca acctttaacg tcggatggcc gcgagacggc acctttaacc
     1321 gagacctcat cacccaggtt aagatcaagg tcttttcacc tggcccgcat ggacacccag
     1381 accaggtccc ctacatcgtg acctgggaag ccttggcttt tgacccccct ccctgggtca
     1441 agccctttgt acaccctaag cctccgcctc ctcttcctcc atccgccccg tctctccccc
     1501 ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta tccagccctc actccttctc
     1561 taggcgccgg aattgaagat ctcgatgagg ccctttcgtc ttcactcgag tttactccct
     1621 atcagtgata gagaacgtat gaagagttta ctccctatca gtgatagaga acgtatgcag
     1681 actttactcc ctatcagtga tagagaacgt ataaggagtt tactccctat cagtgataga
     1741 gaacgtatga ccagtttact ccctatcagt gatagagaac gtatctacag tttactccct
     1801 atcagtgata gagaacgtat atccagttta ctccctatca gtgatagaga acgtatgtcg
     1861 aggtaggcgt gtacggtggg cgcctataaa agcagagctc gtttagtgaa ccgtcagatc
     1921 gcctggagca attccacaac acttttgtct tatacttgga tccgccacca tggcgcctat
     1981 cacggcctac tcccaacaga cgcgaggcct acttggctgc atcatcacta gcctcacagg
     2041 ccgggacagg aaccaggtcg agggggaggt ccaagtggtc tccaccgcaa cacaatcttt
     2101 cctggcgacc tgcgtcaatg gcgtgtgttg gactgtctat catggtgccg gctcaaagac
     2161 ccttgccggc ccaaagggcc caatcaccca aatgtacacc aatgtggacc aggacctcgt
     2221 cggctggcaa gcgccccccg gggcgcgttc cttgacacca tgcacctgcg gcagctcgga
     2281 cctttacttg gtcacgaggc atgccgatgt cattccggtg cgccggcggg gcgacagcag
     2341 ggggagccta ctctccccca ggcccgtctc ctacttgaag ggctcttcgg gcggtccact
     2401 gctctgcccc tcggggcacg ctgtgggcat ctttcgggct gccgtgtgca cccgaggggt
     2461 tgcgaaggcg gtggactttg tacccgtcga gtctatggga accactatgc gntccccggt
     2521 cttcacggac aactcgtccc ctccggccgt accgcagaca ttccaggtgg cccatctaca
     2581 cgcccctact ggtagcggca agagcactaa ggtgccggct gcgtatgcag cccaagggta
     2641 taaggtgctt gtcctgaacc cgtccgtcgc cgccacccta ggtttcgggg cgtatatgtc
     2701 taaggcacat ggtatcgacc ctaacatcag aatcggggta aggaccatca ccacgggtgc
     2761 ccccatcacg tactccacct atggcaagtt tcttgccgac ggtggttgct ctgggggcgc
     2821 ctatgacatc ataatatgtg atgagtgcca ctcaactgac tcgaccacta tcctgggcat
     2881 cggcacagtc ctggaccaag cggagacggc tggagcgcga ctcgtcgtgc tcgccaccgc
     2941 tacgcctccg ggatcggtca ccgtgccaca ttcaaacatc gaggaggtgg ctctgtccag
     3001 cactggagaa atcccctttt atggcaaagc catccccatc gagaccatca agggggggag
     3061 gcacctcatt ttctgccatt ccaagaagaa atgtgatgag ctcgccgcga agctgtccgg
     3121 cctcggactc aatgctgtag catattaccg gggccttgat gtatccgtca taccaactag
     3181 cggagacgtc attgtcgtag caacggacgc tctaatgacg ggctttaccg gtgacttcga
     3241 ctcagtgatc gactgcaata catgtgtcac ccagacagtc gacttcagcc tggacccgac
     3301 cttcaccatt gagacgacga ccgtgccaca agacgcggtg tcacgctcgc agcggcgagg
     3361 caggactggt aggggcagga tgggcattta caggtttgtg actccaggag aacggccctc
     3421 gggcatgttc gattcctcgg ttctgtgcga gtgctatgac gcgggctgtg cttggtacga
     3481 gctcacgccc gccgagacct cagttaggtt gcgggcttac ctaaacacac cagggttgcc
     3541 cgtctgccag gaccatctgg agttctggga gagcgtcttt acaggcctca cccacataga
     3601 cgcccatttc ttgtcccaga ctaagcaggc aggagacaac ttcccctacc tggtagcata
     3661 ccaggctacg gtgtgcgcca gggctcaggc tccacctcca tcgtgggacc aaatgtggaa
     3721 gtgtctcata cggctaaagc ctacgctgca cgggccaacg cccctgctgt ataggctggg
     3781 agccgttcaa aacgaggtta ctaccacaca ccccataacc aaatacatca tggcatgcat
     3841 gtcggctgac ctggaggtcg tcacgagcac ctgggtgctg gtaggcggag tcctagcagc
     3901 tctggccgcg tattgcctga caacaggcag cgtggtcatt gtgggcagga tcatcttgtc
     3961 cggaaagccg gccatcattc ccgacaggga agtcctttac cgggagttcg atgaaatgga
     4021 agagtgctga gcggccgcgc cggcgggccc gaattctacc gggtagggga ggcgcttttc
     4081 ccaaggcagt ctggagcatg cgctttagca gccccgctgg gcacttggcg ctacacaagt
     4141 ggcctctggc ctcgcacaca ttccacatcc accggtaggc gccaaccggc tccgttcttt
     4201 ggtggcccct tcgcgccacc ttctactcct cccctagtca ggaagttccc ccccgccccg
     4261 cagctcgcgt cgtgcaggac gtgacaaatg gaagtagcac gtctcactag tctcgtgcag
     4321 atggacagca ccgctgagca atggaagcgg gtaggccttt ggggcagcgg ccaatagcag
     4381 ctttgctcct tcgctttctg ggctcagagg ctgggaaggg gtgggtccgg gggcgggctc
     4441 aggggcgggc tcaggggcgg ggcgggcgcc cgaaggtcct ccggaggccc ggcattctgc
     4501 acgcttcaaa agcgcacgtc tgccgcgctg ttctcctctt cctcatctcc gggcctttcg
     4561 acctgcagcc caagcttacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg
     4621 acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
     4681 gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
     4741 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
     4801 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
     4861 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
     4921 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc
     4981 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
     5041 ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc
     5101 tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga
     5161 cccgcaagcc cggtgcctga cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg
     5221 cacgacccca tgcatcggcg tctcgagata tcagtggtcc aggctctagt tttgactcaa
     5281 caatatcacc agctgaagcc tatagagtac gagccataga taaaataaaa gattttattt
     5341 agtctccaga aaaagggggg aatgaaagac cccacctgta ggtttggcaa gctagcttaa
     5401 gtaacgccat tttgcaaggc atggaaaaat acataactga gaatagagaa gttcagatca
     5461 aggtcaggaa cagatggaac agggtcgacc ctagagaacc atcagatgtt tccagggtgc
     5521 cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa ccaatcagtt cgcttctcgc
     5581 ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc cctcactcgg
     5641 ggcgccagtc ctccgattga ctgagtcgcc cgggtacccg tgtatccaat aaaccctctt
     5701 gcagttgcat ccgacttgtg gtctcgctgt tccttgggag ggtctcctct gagtgattga
     5761 ctacccgtca gcgggggtct ttcatttggg ggctcgtccg ggatcgggag acccctgccc
     5821 agggaccacc gacccaccac cgggaggtaa gctggctgcc tcgcgcgttt cggtgatgac
     5881 ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat
     5941 gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca
     6001 gccatgaccc agtcacgtag cgatagcgga gtgtagatcc ggctgtggaa tgtgtgtcag
     6061 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     6121 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     6181 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     6241 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     6301 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     6361 ggaggcctag gcttttgcaa aaagcttact ggcttaacta tgcggcatca gagcagattg
     6421 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     6481 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     6541 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     6601 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     6661 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     6721 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     6781 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     6841 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     6901 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     6961 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     7021 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     7081 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     7141 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     7201 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     7261 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     7321 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     7381 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     7441 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     7501 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     7561 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     7621 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     7681 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     7741 ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     7801 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     7861 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     7921 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     7981 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     8041 cggcgtcaac acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     8101 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     8161 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     8221 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     8281 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     8341 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     8401 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     8461 ataaaaatag gcgtatcacg aggccctttc gtcttcaaga attagcttgg ccattgcata
     8521 cgttgtatcc atatcataat atgtacattt atattggctc atgtccaaca ttaccgccat
     8581 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
     8641 gcccatatat ggagt
//
