LOCUS       sequence-487710-        7539 bp    DNA     circular SYN 02-OCT-2025
DEFINITION  Cre-dependent AAV vector for cocaine-gated chemogenetic anion 
            channel.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7539)
  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 7539)
  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-GlyR-IRES-mCherry
FEATURES             Location/Qualifiers
     source          1..7539
                     /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)."
     CDS             3270..3296
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    join(3522..4096,4097..4099,4100..4108)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3690..3707)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             4121..4828
                     /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(4263..4281)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4531..4550
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(4841..4874)
                     /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(4937..4970)
                     /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(4989..5008)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          4991..5123
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     repeat_region   5722..5851
                     /label=AAV2 ITR
     rep_origin      5923..6378
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6010..6029)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6220..6241
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6404..6508
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6509..6577,6578..7369)
                     /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(6727..6746)
                     /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 tcgctagcaa gcttgaattc gccaccatgc
     2161 gctgttctcc aggcggcgtg tggctcgccc tggctgcttc ccttctgcac gttagcctgc
     2221 agggtgagtt ccagcgcaaa ctgtataagg agcttgttaa gaattataac cccctggagc
     2281 ggccggtcgc aaatgattcc cagccactga cagtgtactt cagcctctcc ttgctgcaga
     2341 tcatggacgt ggatgaaaag aaccaggtgc tgaccactaa tatttggttg cagatgtcct
     2401 ggaccgatca ctacttgcag tggaatgtga gcgaataccc aggtgtaaag actgtaagat
     2461 tccctgacgg ccaaatctgg aaaccagata tcctgctgta caacagcgca gacgaaaggt
     2521 ttgatgcaac atttcacacc aacgtgttgg tcaattcttc aggccactgc cagtacggac
     2581 cccctggaat cttcaagtcc tcatgctata tcgacgtccg ctggtttccc ttcgacgtcc
     2641 agcactgcaa actcaaattc gggagctgga gctacggcaa atggagcctg gatctgcaaa
     2701 tgcaggaggc tgacatctct ggttacatcc cgaatgggga gtgggacctt gtgggaatcc
     2761 ccggtaaaag aagcgagcga ttttatgaat gctgcaaaga gcccttccca gatgtcacct
     2821 tcacagtgac catgcggaga cgcatgggtt attatctgat ccaaatgtat atcccaaact
     2881 tgcttatagt gattttgtca tggatctcct tctggattaa tatggacgcc gctccagcta
     2941 gggtcggact gggcatcacc acagtgctga caatgactac tcagagctca ggcagccgag
     3001 ccagcttgcc caaggtttct tacgtgaagg ccatcgatat ctggatggct gtctgccttc
     3061 tgtttgtctt cagcgcactg ctggaatacg ccgctgtcaa ttttgtgtct cgacagcata
     3121 aagagctgtt gcggttcaga agaaaacgac gccaccacaa agaggatgag gcaggagaag
     3181 gacgcttcaa ctttagcgcc tatggtatgg gacctgcttg cctccaggct aaagacggaa
     3241 tttccgtgaa gggagccaac aatagcaact acccttatga tgtcccagac tatgcaacaa
     3301 ccaacccacc ccctgctcca tctaagagcc cggaggaaat gcgcaaactc tttattcaga
     3361 gagcgaaaaa gatcgacaaa atctcccgga tcggattccc catggctttc ctgattttca
     3421 acatgtttta ttggatcatc tacaagattg tgcgaaggga ggacgtacac aaccagtaac
     3481 tcgagtctag agcggccgca attcccccmc cccccccccc cccccyctcc ctcccccccc
     3541 ccctaacgtt actggccgaa gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt
     3601 attttccacc atattgccgt cttttggcaa tgtgagggcc cggaaacctg gccctgtctt
     3661 cttgacgagc attcctaggg gtctttcccc tctcgccaaa ggaatgcaag gtctgttgaa
     3721 tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac
     3781 cctttgcagg cagcggaacc ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg
     3841 tgtataagat acacctgcaa aggcggcaca accccagtgc cacgttgtga gttggatagt
     3901 tgtggaaaga gtcaaatggc tctcctaagc gtattcaaca aggggctgaa ggatgcccag
     3961 aaggtacccc attgtatggg atctgatctg gggcctcggt gcacatgctt tacatgtgtt
     4021 tagtcgaggt taaaaaaacg tctaggcccc ccgaaccacg gggacgtggt tttcctttga
     4081 aaaacacgat gataatatgg ccacaaccat gggggatccg gtgagcaagg gcgaggagga
     4141 taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa
     4201 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
     4261 cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc
     4321 tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt
     4381 gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
     4441 cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa
     4501 gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg
     4561 ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca
     4621 gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc
     4681 caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc
     4741 ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac
     4801 cggcggcatg gacgagctgt acaagtaatc tagcaagctt ataacttcgt atagcataca
     4861 ttatacgaag ttatgtgtcg atggtgatgc ttggcaattc gggtcgatgg tgaagcattg
     4921 gtcttcctat tttgatataa cttcgtatag gatactttat acgaagttat ggggggcgcg
     4981 ccatcatcac ctgtggagag aaaggcaaag tggatgtcag taagaccaat aggtgcctat
     5041 cagaaacgca agagtcttct ctgtctcgac aagcccagtt tctattggtc tccttaaacc
     5101 tgtcttgtaa ccttgatact tacctgatcc aacggctgcg acttggggca gggggtccta
     5161 ggggtggtct ggccaggagc cgcgggggcg gactgcgcgg tgcccaggag cacagctgcg
     5221 ctctcaggca cgacacgact cctccgctgc ccaccgcaga ctgaggcagc gctgagtcgc
     5281 cggcgccgca gcgcagatgg tcgcgcccgt gcccccctat ctcgcgcctc gcgtggtgcg
     5341 gtccggctgg gccggcggcg gcgcggacgc gaccaaggtg gccgggaagg ggagtttgcg
     5401 ggggaccggc gagtgacgtc agcgcgcctt cagtgctgag gcggcggtgg cgcgcgccgc
     5461 caggcggggg cgaaggcact gtccgcggtg ctgaagctgg cagtgcgcac gcgcctcgcc
     5521 gcatcctgtt tcccctcccc ctctctgata ggggatgcgc aatttgggga atgggggttg
     5581 ggtgcttgtc cagtgggtcg gggtcggtcg tcaagcatgc tttaaatact gaaaagggcc
     5641 aattccagca cactggagct cggatcttcc tagagcatgg ctacgtagat aagtagcatg
     5701 gcgggttaat cattaactac aaggaacccc tagtgatgga gttggccact ccctctctgc
     5761 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     5821 gggcggcctc agtgagcgag cgagcgcgca gctggcgtaa tagcgaagag gcccgcaccg
     5881 atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg
     5941 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     6001 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     6061 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     6121 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     6181 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     6241 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     6301 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     6361 tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     6421 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     6481 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     6541 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     6601 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     6661 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     6721 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     6781 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     6841 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     6901 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     6961 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     7021 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     7081 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     7141 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     7201 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     7261 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     7321 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     7381 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     7441 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     7501 ctgagcgtca gaccccgtag aaaagatcaa aggatcttc
//
