LOCUS       sequence-366445-        8276 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Doxycycline-dependent expression of human cGAS gene in mammalian 
            cells by retroviral transduction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8276)
  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 8276)
  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-FLAG-human cGAS N-T2A-deltaN-HA
FEATURES             Location/Qualifiers
     source          1..8276
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        311..514
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             515..690
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    753..952
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1153..1569
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1499..1521
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1537..1553
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        1609..1958
                     /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    1617..1635
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1653..1671
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1689..1707
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1725..1743
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1761..1779
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1797..1815
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1833..1851
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1897..1921
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1965..1974
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1974..1997
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2481..2534
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3624..3650
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        3684..4183
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3719..3742)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     4116..4135
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             4204..4803
                     /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(4204..4223)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4700..4720
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4984..5409
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(5512..5534)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5670..5690)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5675..6004
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5855..5990
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6026..6045
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6126..6143)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6297..6885)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6377..6396)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7056..7847,7848..7916))
                     /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     7679..7698
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7917..8021)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8089..8107
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(8207..8276,1..310)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
       61 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
      121 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
      181 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
      241 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
      301 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
      361 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
      421 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
      481 gcgtgtacgg tgggaggtct atataagcag agctcaataa aagagcccac aacccctcac
      541 tcggcgcgcc agtcttccga tagactgcgt cgcccgggta cccgtattcc caataaagcc
      601 tcttgctgtt tgcatccgaa tcgtggtctc gctgttcctt gggagggtct cctctgagtg
      661 attgactacc cacgacgggg gtctttcatt tgggggctcg tccgggattt ggagacccct
      721 gcccagggac caccgaccca ccaccgggag gtaagctggc cagcaactta tctgtgtctg
      781 tccgattgtc tagtgtctat gtttgatgtt atgcgcctgc gtctgtacta gttagctaac
      841 tagctctgta tctggcggac ccgtggtgga actgacgagt tctgaacacc cggccgcaac
      901 cctgggagac gtcccaggga ctttgggggc cgtttttgtg gcccgacctg aggaagggag
      961 tcgatgtgga atccgacccc gtcaggatat gtggttctgg taggagacga gaacctaaaa
     1021 cagttcccgc ctccgtctga atttttgctt tcggtttgga accgaagccg cgcgtcttgt
     1081 ctgctgcagc gctgcagcat cgttctgtgt tgtctctgtc tgactgtgtt tctgtatttg
     1141 tctgaaaatt agggccagac tgttaccact cccttaagtt tgaccttagg tcactggaaa
     1201 gatgtcgagc ggatcgctca caaccagtcg gtagatgtca agaagagacg ttgggttacc
     1261 ttctgctctg cagaatggcc aacctttaac gtcggatggc cgcgagacgg cacctttaac
     1321 cgagacctca tcacccaggt taagatcaag gtcttttcac ctggcccgca tggacaccca
     1381 gaccaggtcc cctacatcgt gacctgggaa gccttggctt ttgacccccc tccctgggtc
     1441 aagccctttg tacaccctaa gcctccgcct cctcttcctc catccgcccc gtctctcccc
     1501 cttgaacctc ctcgttcgac cccgcctcga tcctcccttt atccagccct cactccttct
     1561 ctaggcgccg gaattgaaga tctcgatgag gccctttcgt cttcactcga gtttactccc
     1621 tatcagtgat agagaacgta tgaagagttt actccctatc agtgatagag aacgtatgca
     1681 gactttactc cctatcagtg atagagaacg tataaggagt ttactcccta tcagtgatag
     1741 agaacgtatg accagtttac tccctatcag tgatagagaa cgtatctaca gtttactccc
     1801 tatcagtgat agagaacgta tatccagttt actccctatc agtgatagag aacgtatgtc
     1861 gaggtaggcg tgtacggtgg gcgcctataa aagcagagct cgtttagtga accgtcagat
     1921 cgcctggagc aattccacaa cacttttgtc ttatacttgg atccgccacc atggattaca
     1981 aggatgacga cgataagcag ccttggcacg gaaaggccat gcagagagct tccgaggccg
     2041 gagccactgc ccccaaggct tccgcacgga atgccagggg cgccccgatg gatcccaccg
     2101 agtctccggc tgcccccgag gccgccctgc ctaaggcggg aaagttcggc cccgccagga
     2161 agtcgggatc ccggcagaaa aagagcgccc cggacaccca ggagaggccg cccgtccgcg
     2221 caactggggc ccgcgccaaa aaggcccctc agcgcgccca ggacacgcag ccgtctgacg
     2281 ccaccagcgc ccctggggca gaggggctgg agcctcctgc ggctcgggag ccggctcttt
     2341 ccagggctgg ttcttgccgc cagaggggcg cgcgctgctc cacgaagcca agacctccgc
     2401 ccgggccctg ggacgtgccc agccccggcc tgccggtctc ggcccccatt ctcgtacgga
     2461 gggatgcggc gggtagcgga gagggcagag gcagtctgct gacatgcggt gacgtggaag
     2521 agaatcccgg ccctcctggg gcctcgaagc tccgggcggt tttggagaag ttgaagctca
     2581 gccgcgatga tatctccacg gcggcgggga tggtgaaagg ggttgtggac cacctgctgc
     2641 tcagactgaa gtgcgactcc gcgttcagag gcgtcgggct gctgaacacc gggagctact
     2701 atgagcacgt gaagatttct gcacctaatg aatttgatgt catgtttaaa ctggaagtcc
     2761 ccagaattca actagaagaa tattccaaca ctcgtgcata ttactttgtg aaatttaaaa
     2821 gaaatccgaa agaaaatcct ctgagtcagt ttttagaagg tgaaatatta tcagcttcta
     2881 agatgctgtc aaagtttagg aaaatcatta aggaagaaat taacgacatt aaagatacag
     2941 atgtcatcat gaagaggaaa agaggaggga gccctgctgt aacacttctt attagtgaaa
     3001 aaatatctgt ggatataacc ctggctttgg aatcaaaaag tagctggcct gctagcaccc
     3061 aagaaggcct gcgcattcaa aactggcttt cagcaaaagt taggaagcaa ctacgactaa
     3121 agccatttta ccttgtaccc aagcatgcaa aggaaggaaa tggtttccaa gaagaaacat
     3181 ggcggctatc cttctctcac atcgaaaagg aaattttgaa caatcatgga aaatctaaaa
     3241 cgtgctgtga aaacaaagaa gagaaatgtt gcaggaaaga ttgtttaaaa ctaatgaaat
     3301 accttttaga acagctgaaa gaaaggttta aagacaaaaa acatctggat aaattctctt
     3361 cttatcatgt gaaaactgcc ttctttcacg tatgtaccca gaaccctcaa gacagtcagt
     3421 gggaccgcaa agacctgggc ctctgctttg ataactgcgt gacatacttt cttcagtgcc
     3481 tcaggacaga aaaacttgag aattatttta ttcctgaatt caatctattc tctagcaact
     3541 taattgacaa aagaagtaaa gaatttctga caaagcaaat tgaatatgaa agaaacaatg
     3601 agtttccagt ttttgatgaa ttttacccat acgatgttcc agattacgct tgagcggccg
     3661 cgccggcggg cccgaattct accgggtagg ggaggcgctt ttcccaaggc agtctggagc
     3721 atgcgcttta gcagccccgc tgggcacttg gcgctacaca agtggcctct ggcctcgcac
     3781 acattccaca tccaccggta ggcgccaacc ggctccgttc tttggtggcc ccttcgcgcc
     3841 accttctact cctcccctag tcaggaagtt cccccccgcc ccgcagctcg cgtcgtgcag
     3901 gacgtgacaa atggaagtag cacgtctcac tagtctcgtg cagatggaca gcaccgctga
     3961 gcaatggaag cgggtaggcc tttggggcag cggccaatag cagctttgct ccttcgcttt
     4021 ctgggctcag aggctgggaa ggggtgggtc cgggggcggg ctcaggggcg ggctcagggg
     4081 cggggcgggc gcccgaaggt cctccggagg cccggcattc tgcacgcttc aaaagcgcac
     4141 gtctgccgcg ctgttctcct cttcctcatc tccgggcctt tcgacctgca gcccaagctt
     4201 accatgaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc
     4261 gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg
     4321 gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc
     4381 gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg
     4441 gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc
     4501 ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag
     4561 gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg
     4621 ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc
     4681 ctggagacct ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc
     4741 gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc
     4801 tgacgcccgc cccacgaccc gcagcgcccg accgaaagga gcgcacgacc ccatgcatcg
     4861 gcgtctcgag atatcagtgg tccaggctct agttttgact caacaatatc accagctgaa
     4921 gcctatagag tacgagccat agataaaata aaagatttta tttagtctcc agaaaaaggg
     4981 gggaatgaaa gaccccacct gtaggtttgg caagctagct taagtaacgc cattttgcaa
     5041 ggcatggaaa aatacataac tgagaataga gaagttcaga tcaaggtcag gaacagatgg
     5101 aacagggtcg accctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg
     5161 accctgtgcc ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc
     5221 tgctccccga gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat
     5281 tgactgagtc gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt
     5341 gtggtctcgc tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg
     5401 tctttcattt gggggctcgt ccgggatcgg gagacccctg cccagggacc accgacccac
     5461 caccgggagg taagctggct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     5521 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     5581 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg
     5641 tagcgatagc ggagtgtaga tccggctgtg gaatgtgtgt cagttagggt gtggaaagtc
     5701 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     5761 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     5821 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     5881 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     5941 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     6001 caaaaagctt actggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     6061 atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag gcgctcttcc
     6121 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     6181 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     6241 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     6301 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     6361 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     6421 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     6481 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     6541 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     6601 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     6661 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     6721 tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     6781 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     6841 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     6901 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     6961 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     7021 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     7081 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     7141 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     7201 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     7261 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     7321 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tgcaggcatc
     7381 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     7441 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     7501 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     7561 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     7621 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aacacgggat
     7681 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     7741 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     7801 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     7861 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     7921 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     7981 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     8041 ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc
     8101 acgaggccct ttcgtcttca agaattagct tggccattgc atacgttgta tccatatcat
     8161 aatatgtaca tttatattgg ctcatgtcca acattaccgc catgttgaca ttgattattg
     8221 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatgga
//
