LOCUS       sequence-366485-        7703 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Doxycycline-dependent expression of human cGAS gene (with R255E 
            mutation) in mammalian cells by retroviral transduction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7703)
  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 7703)
  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-human cGASdeltaN R255E-HA
FEATURES             Location/Qualifiers
     source          1..7703
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        329..532
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             533..708
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    771..970
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1171..1587
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1517..1539
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1555..1571
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        1627..1976
                     /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    1635..1653
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1671..1689
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1707..1725
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1743..1761
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1779..1797
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1815..1833
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1851..1869
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1915..1939
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             3081..3107
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        3141..3628
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3176..3199)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3561..3580
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3649..4248
                     /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(3649..3668)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4145..4165
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4429..4854
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(4957..4979)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5115..5135)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5120..5449
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5300..5435
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5471..5490
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5571..5588)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5742..6330)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5822..5841)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6501..7292,7293..7361))
                     /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     7124..7143
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7362..7466)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7534..7552
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(7652..7703,1..328)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
       61 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      121 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      181 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      241 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      301 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
      361 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      421 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
      481 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctcaataaaa
      541 gagcccacaa cccctcactc ggcgcgccag tcttccgata gactgcgtcg cccgggtacc
      601 cgtattccca ataaagcctc ttgctgtttg catccgaatc gtggtctcgc tgttccttgg
      661 gagggtctcc tctgagtgat tgactaccca cgacgggggt ctttcatttg ggggctcgtc
      721 cgggatttgg agacccctgc ccagggacca ccgacccacc accgggaggt aagctggcca
      781 gcaacttatc tgtgtctgtc cgattgtcta gtgtctatgt ttgatgttat gcgcctgcgt
      841 ctgtactagt tagctaacta gctctgtatc tggcggaccc gtggtggaac tgacgagttc
      901 tgaacacccg gccgcaaccc tgggagacgt cccagggact ttgggggccg tttttgtggc
      961 ccgacctgag gaagggagtc gatgtggaat ccgaccccgt caggatatgt ggttctggta
     1021 ggagacgaga acctaaaaca gttcccgcct ccgtctgaat ttttgctttc ggtttggaac
     1081 cgaagccgcg cgtcttgtct gctgcagcgc tgcagcatcg ttctgtgttg tctctgtctg
     1141 actgtgtttc tgtatttgtc tgaaaattag ggccagactg ttaccactcc cttaagtttg
     1201 accttaggtc actggaaaga tgtcgagcgg atcgctcaca accagtcggt agatgtcaag
     1261 aagagacgtt gggttacctt ctgctctgca gaatggccaa cctttaacgt cggatggccg
     1321 cgagacggca cctttaaccg agacctcatc acccaggtta agatcaaggt cttttcacct
     1381 ggcccgcatg gacacccaga ccaggtcccc tacatcgtga cctgggaagc cttggctttt
     1441 gacccccctc cctgggtcaa gccctttgta caccctaagc ctccgcctcc tcttcctcca
     1501 tccgccccgt ctctccccct tgaacctcct cgttcgaccc cgcctcgatc ctccctttat
     1561 ccagccctca ctccttctct aggcgccgga attgaagatc tcgatgaggc cctttcgtct
     1621 tcactcgagt ttactcccta tcagtgatag agaacgtatg aagagtttac tccctatcag
     1681 tgatagagaa cgtatgcaga ctttactccc tatcagtgat agagaacgta taaggagttt
     1741 actccctatc agtgatagag aacgtatgac cagtttactc cctatcagtg atagagaacg
     1801 tatctacagt ttactcccta tcagtgatag agaacgtata tccagtttac tccctatcag
     1861 tgatagagaa cgtatgtcga ggtaggcgtg tacggtgggc gcctataaaa gcagagctcg
     1921 tttagtgaac cgtcagatcg cctggagcaa ttccacaaca cttttgtctt atacttggat
     1981 ccgccaccat gcctggggcc tcgaagctcc gggcggtttt ggagaagttg aagctcagcc
     2041 gcgatgatat ctccacggcg gcggggatgg tgaaaggggt tgtggaccac ctgctgctca
     2101 gactgaagtg cgactccgcg ttcagaggcg tcgggctgct gaacaccggg agctactatg
     2161 agcacgtgaa gatttctgca cctaatgaat ttgatgtcat gtttaaactg gaagtcccca
     2221 gaattcaact agaagaatat tccaacactc gtgcatatta ctttgtgaaa tttaaagaaa
     2281 atccgaaaga aaatcctctg agtcagtttt tagaaggtga aatattatca gcttctaaga
     2341 tgctgtcaaa gtttaggaaa atcattaagg aagaaattaa cgacattaaa gatacagatg
     2401 tcatcatgaa gaggaaaaga ggagggagcc ctgctgtaac acttcttatt agtgaaaaaa
     2461 tatctgtgga tataaccctg gctttggaat caaaaagtag ctggcctgct agcacccaag
     2521 aaggcctgcg cattcaaaac tggctttcag caaaagttag gaagcaacta cgactaaagc
     2581 cattttacct tgtacccaag catgcaaagg aaggaaatgg tttccaagaa gaaacatggc
     2641 ggctatcctt ctctcacatc gaaaaggaaa ttttgaacaa tcatggaaaa tctaaaacgt
     2701 gctgtgaaaa caaagaagag aaatgttgca ggaaagattg tttaaaacta atgaaatacc
     2761 ttttagaaca gctgaaagaa aggtttaaag acaaaaaaca tctggataaa ttctcttctt
     2821 atcatgtgaa aactgccttc tttcacgtat gtacccagaa ccctcaagac agtcagtggg
     2881 accgcaaaga cctgggcctc tgctttgata actgcgtgac atactttctt cagtgcctca
     2941 ggacagaaaa acttgagaat tattttattc ctgaattcaa tctattctct agcaacttaa
     3001 ttgacaaaag aagtaaagaa tttctgacaa agcaaattga atatgaaaga aacaatgagt
     3061 ttccagtttt tgatgaattt tacccatacg atgttccaga ttacgcttga gcggccgcgc
     3121 cggcgggccc gaattctacc gggtagggga ggcgcttttc ccaaggcagt ctggagcatg
     3181 cgctttagca gccccgctgg gcacttggcg ctacacaagt ggcctctggc ctcgcacaca
     3241 ttccacatcc accggtaggc gccaaccggc tccgttcttt ggtggcccct tcgcgccacc
     3301 ttctactcct cccctagtca ggaagttccc ccccgccccg cagctcgcgt cgtgcaggac
     3361 gtgacaaatg gaagtagcac gtctcactag tctcgtgcag atggacagca ccgctgagca
     3421 atggaagcgg gtaggccttt ggggcagcgg ccaatagcag ctttgctcct tcgctttctg
     3481 ggctcagagg ctgggaaggg gtgggtccgg gggcgggctc aggggcgggg cgggcgcccg
     3541 aaggtcctcc ggaggcccgg cattctgcac gcttcaaaag cgcacgtctg ccgcgctgtt
     3601 ctcctcttcc tcatctccgg gcctttcgac ctgcagccca agcttaccat gaccgagtac
     3661 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
     3721 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
     3781 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     3841 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     3901 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     3961 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     4021 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     4081 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
     4141 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     4201 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg cccgccccac
     4261 gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcggcgtc tcgagatatc
     4321 agtggtccag gctctagttt tgactcaaca atatcaccag ctgaagccta tagagtacga
     4381 gccatagata aaataaaaga ttttatttag tctccagaaa aaggggggaa tgaaagaccc
     4441 cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat ggaaaaatac
     4501 ataactgaga atagagaagt tcagatcaag gtcaggaaca gatggaacag ggtcgaccct
     4561 agagaaccat cagatgtttc cagggtgccc caaggacctg aaatgaccct gtgccttatt
     4621 tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc cccgagctca
     4681 ataaaagagc ccacaacccc tcactcgggg cgccagtcct ccgattgact gagtcgcccg
     4741 ggtacccgtg tatccaataa accctcttgc agttgcatcc gacttgtggt ctcgctgttc
     4801 cttgggaggg tctcctctga gtgattgact acccgtcagc gggggtcttt catttggggg
     4861 ctcgtccggg atcgggagac ccctgcccag ggaccaccga cccaccaccg ggaggtaagc
     4921 tggctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     4981 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     5041 agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt
     5101 gtagatccgg ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     5161 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     5221 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     5281 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     5341 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     5401 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agcttactgg
     5461 cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata
     5521 ccgcacagat gcgtaaggag aaaataccgc atcaggcgct cttccgcttc ctcgctcact
     5581 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5641 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5701 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5761 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5821 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5881 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5941 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     6001 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     6061 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     6121 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     6181 aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     6241 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     6301 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     6361 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     6421 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     6481 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6541 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6601 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6661 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6721 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6781 ccagttaata gtttgcgcaa cgttgttgcc attgctgcag gcatcgtggt gtcacgctcg
     6841 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6901 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6961 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     7021 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     7081 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaacac gggataatac cgcgccacat
     7141 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     7201 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     7261 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     7321 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     7381 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     7441 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa
     7501 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt
     7561 cttcaagaat tagcttggcc attgcatacg ttgtatccat atcataatat gtacatttat
     7621 attggctcat gtccaacatt accgccatgt tgacattgat tattgactag ttattaatag
     7681 taatcaatta cggggtcatt agt
//
