LOCUS       sequence-487707-        7614 bp    DNA     circular SYN 02-OCT-2025
DEFINITION  Cre-dependent AAV vector for cocaine-gated chemogenetic cation 
            channel.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7614)
  AUTHORS   Gomez JL, Magnus CJ, Bonaventura J, Solis O, Curry FP, Levinstein 
            MR, Budinich RC, Carlton ML, Ventriglia EN, Lam S, Wang L, 
            Schoenborn I, Dunne W, Michaelides M, Sternson SM
  TITLE     Cocaine chemogenetics blunts drug-seeking by synthetic physiology.
  JOURNAL   Nature. 2025 Aug 27. doi: 10.1038/s41586-025-09427-8.
  PUBMED    40866713
REFERENCE   2  (bases 1 to 7614)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 2, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2025 Aug 27. doi: 10.1038/s41586-025-09427-8."
COMMENT     Sequence Label: AAV Syn-FLEX-coca-5HT3-IRES-mCherry
FEATURES             Location/Qualifiers
     source          1..7614
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   826..955
                     /label=AAV2 ITR
     polyA_signal    1117..1238
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1154..1173)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1208..1227
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    1315..1903
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1421..1432
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1830..1850
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    1971..2004
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2075..2108
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     misc_feature    join(3599..4171,4172..4174,4175..4183)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3765..3782)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             4196..4903
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(4338..4356)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4606..4625
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(4916..4949)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(5012..5045)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     primer_bind     complement(5064..5083)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          5066..5198
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     repeat_region   5797..5926
                     /label=AAV2 ITR
     rep_origin      5998..6453
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6085..6104)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6295..6316
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6479..6583
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6584..6652,6653..7444)
                     /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     complement(6802..6821)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctgcg cgctcgctcg
      841 ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg cccggcctca
      901 gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg ttccttgtag
      961 ttaatgatta acccgccatg ctacttatct acgtagccat gctctaggaa gatccttatc
     1021 gattttacca catttgtaga ggttttactt gctttaaaaa acctcccaca tctccccctg
     1081 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     1141 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     1201 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgctc gaagcggccg
     1261 gccgcccggg cgacctagat gcattcgacg gccgcccggg tcgacggtat cgatgcgggg
     1321 aggcggccca aagggagatc cgactcgtct gagggcgaag gcgaagacgc ggaagaggcc
     1381 gcagagccgg cagcaggccg cgggaaggaa ggtccgctgg attgagggcc gaagggacgt
     1441 agcagaagga cgtcccgcgc agaatccagg tggcaacaca ggcgagcagc caaggaaagg
     1501 acgatgattt ccccgacaac accacggaat tgtcagtgcc caacagccga gcccctgtcc
     1561 agcagcgggc aaggcaggcg gcgatgagtt ccgccgtggc aatagggagg gggaaagcga
     1621 aagtcccgga aaggagctga caggtggtgg caatgcccca accagtgggg gttgcgtcag
     1681 caaacacagt gcacaccacg ccacgttgcc tgacaacggg ccacaactcc tcataaagag
     1741 acagcaacca ggatttatac aaggaggaga aaatgaaagc catacgggaa gcaatagcat
     1801 gatacaaagg cattaaagca gcgtatccac atagcgtaaa aggagcaaca tagttaagaa
     1861 taccagtcaa tctttcacaa attttgtaat ccagaggttg attcttaaga gcttcgagtg
     1921 gatcatctgc aaattcgccc ttttcagtgt ttcgaacctc ttcgagggac ataacttcgt
     1981 ataatgtatg ctatacgaag ttatgggtcg atggtgagat ctggactaga gggtcgatgg
     2041 tgatgcttgg atcaaaatag gaagaatccc aatgataact tcgtataaag tatcctatac
     2101 gaagttatac tagtcatatg ggatccgaat tcgctagcga attcgccacc atgcgctgtt
     2161 ctccaggcgg cgtgtggctc gccctggctg cttcccttct gcacgttagc ctgcagggtg
     2221 agttccagcg caaactgtat aaggagcttg ttaagaatta taaccccctg gagcggccgg
     2281 tcgcaaatga ttcccagcca ctgacagtgt acttcagcct ctccttgctg cagatcatgg
     2341 acgtggatga aaagaaccag gtgctgacca ctaatatttg gttgcagatg tcctggaccg
     2401 atcactactt gcagtggaat gtgagcgaat acccaggtgt aaagactgta agattccctg
     2461 acggccaaat ctggaaacca gatatcctgc tgtacaacag cgcagacgaa aggtttgatg
     2521 caacatttca caccaacgtg ttggtcaatt cttcaggcca ctgccagtac ggaccccctg
     2581 gaatcttcaa gtcctcatgc tatatcgacg tccgctggtt tcccttcgac gtccagcact
     2641 gcaaactcaa attcgggagc tggagctacg gcaaatggag cctggatctg caaatgcagg
     2701 aggctgacat ctctggttac atcccgaatg gggagtggga ccttgtggga atccccggta
     2761 aaagaagcga gcgatttttt gaatgctgca aggaaccctt ccctgacgta acattcacag
     2821 ttatcatcag aagaaggcca ttgttctacg ccgttagttt gttgctcccc agtatttttc
     2881 tcatggtcgt ggacatcgtg ggattttgtc tcccacctga tagcggggag agggtctcct
     2941 ttaagattac cttgttgctc ggctattctg tatttctgat catcgtgtcc gatacccttc
     3001 ctgccacaat cggcactccg ctgataggag tgtatttcgt cgtgtgtatg gcactcctgg
     3061 tgataagtct ggcggaaact atcttcattg tacggctggt acataagcag gacctgcaaa
     3121 gacccgtgcc agactggttg cgacaccttg tgctggacag aattgcatgg attctgtgtc
     3181 ttggcgagca acctatggcc caccggccac ctgcaacctt tcaagccaac aagacagacg
     3241 attgtagtgg gtctgatctg ttgcctgcta tggggaatca ctgctcccat gttgggggac
     3301 cacaagattt ggaaaagacc ccacgggggc ggggatcacc ccttcctcct ccccgagaag
     3361 cctctctcgc tgtccggggg ctgctccagg aactgtcaag catccgacat tttctggaga
     3421 agcgggacga gatgcgcgaa gtcgctcgcg actggctgcg agtgggctac gtccttgaca
     3481 ggctgctgtt tcggatctac ttgctggcgg tgctggctta ttccattact ctggtgacac
     3541 tctggtccat atggcactac agttaggcgg ccgcaattsc csccsycccc cccccccscc
     3601 cctcyccccc ccccccccta acgttactgg ccgaagccgc ttggaataag gccggtgtgc
     3661 gtttgtctat atgttatttt ccaccatatt gccgtctttt ggcaatgtga gggcccggaa
     3721 acctggccct gtcttcttga cgagcattcc taggggtctt tcccctctcg ccaaaggaat
     3781 gcaaggtctg ttgaatgtcg tgaaggaagc agttcctctg gaagcttctt gaagacaaac
     3841 aacgtctgta gcgacccttt gcaggcagcg gaacccccca cctggcgaca ggtgcctctg
     3901 cggccaaaag ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc agtgccacgt
     3961 tgtgagttgg atagttgtgg aaagagtcaa atggctctcc taagcgtatt caacaagggg
     4021 ctgaaggatg cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca
     4081 tgctttacat gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa ccacggggac
     4141 gtggttttcc tttgaaaaac acgatgataa tatggccaca accatggggg atccggtgag
     4201 caagggcgag gaggataaca tggccatcat caaggagttc atgcgcttca aggtgcacat
     4261 ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta
     4321 cgagggcacc cagaccgcca agctgaaggt gaccaagggt ggccccctgc ccttcgcctg
     4381 ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc tacgtgaagc accccgccga
     4441 catccccgac tacttgaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa
     4501 cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcgagtt
     4561 catctacaag gtgaagctgc gcggcaccaa cttcccctcc gacggccccg taatgcagaa
     4621 gaagaccatg ggctgggagg cctcctccga gcggatgtac cccgaggacg gcgccctgaa
     4681 gggcgagatc aagcagaggc tgaagctgaa ggacggcggc cactacgacg ctgaggtcaa
     4741 gaccacctac aaggccaaga agcccgtgca gctgcccggc gcctacaacg tcaacatcaa
     4801 gttggacatc acctcccaca acgaggacta caccatcgtg gaacagtacg aacgcgccga
     4861 gggccgccac tccaccggcg gcatggacga gctgtacaag taatctagca agcttataac
     4921 ttcgtatagc atacattata cgaagttatg tgtcgatggt gatgcttggc aattcgggtc
     4981 gatggtgaag cattggtctt cctattttga tataacttcg tataggatac tttatacgaa
     5041 gttatggggg gcgcgccatc atcacctgtg gagagaaagg caaagtggat gtcagtaaga
     5101 ccaataggtg cctatcagaa acgcaagagt cttctctgtc tcgacaagcc cagtttctat
     5161 tggtctcctt aaacctgtct tgtaaccttg atacttacct gatccaacgg ctgcgacttg
     5221 gggcaggggg tcctaggggt ggtctggcca ggagccgcgg gggcggactg cgcggtgccc
     5281 aggagcacag ctgcgctctc aggcacgaca cgactcctcc gctgcccacc gcagactgag
     5341 gcagcgctga gtcgccggcg ccgcagcgca gatggtcgcg cccgtgcccc cctatctcgc
     5401 gcctcgcgtg gtgcggtccg gctgggccgg cggcggcgcg gacgcgacca aggtggccgg
     5461 gaaggggagt ttgcggggga ccggcgagtg acgtcagcgc gccttcagtg ctgaggcggc
     5521 ggtggcgcgc gccgccaggc gggggcgaag gcactgtccg cggtgctgaa gctggcagtg
     5581 cgcacgcgcc tcgccgcatc ctgtttcccc tccccctctc tgatagggga tgcgcaattt
     5641 ggggaatggg ggttgggtgc ttgtccagtg ggtcggggtc ggtcgtcaag catgctttaa
     5701 atactgaaaa gggccaattc cagcacactg gagctcggat cttcctagag catggctacg
     5761 tagataagta gcatggcggg ttaatcatta actacaagga acccctagtg atggagttgg
     5821 ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac
     5881 gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgg cgtaatagcg
     5941 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     6001 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     6061 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     6121 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     6181 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     6241 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     6301 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     6361 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     6421 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     6481 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     6541 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     6601 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     6661 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     6721 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     6781 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     6841 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     6901 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     6961 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     7021 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     7081 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     7141 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     7201 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     7261 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     7321 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     7381 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     7441 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     7501 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     7561 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttc
//
