LOCUS       sequence-333368-        7122 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV plasmid containing a mKate2 and a light-gated adenylate cyclase 
            biPAC.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7122)
  AUTHORS   Zhang SX, Lutas A, Yang S, Diaz A, Fluhr H, Nagel G, Gao S, 
            Andermann ML
  TITLE     Hypothalamic dopamine neurons motivate mating through persistent 
            cAMP signalling.
  JOURNAL   Nature. 2021 Sep;597(7875):245-249. doi: 10.1038/s41586-021-03845-0.
            Epub 2021 Aug 25.
  PUBMED    34433964
REFERENCE   2  (bases 1 to 7122)
  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"; doi: 
            "10.1038/s41586-021-03845-0"; journalName: "Nature"; date: 
            "2021-09"; volume: "597"; issue: "7875"; pages: "245-249"
COMMENT     Sequence Label: pAAV-DIO-EF1-alpha-mKate2-biPAC
FEATURES             Location/Qualifiers
     source          1..7122
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(109..697)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(189..208)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(868..1659,1660..1728))
                     /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     1491..1510
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1729..1833)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1901..1919
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1957..1979)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2079..2098
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(2115..2570)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2252..2273)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2464..2483
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2645..2785
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2656..2785
                     /label=AAV2 ITR
     polyA_signal    2903..3014
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     misc_feature    3048..3636
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3154..3165
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     3563..3583
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(3661..3694)
                     /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(3745..3778)
                     /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)."
     regulatory      3792..3801
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3798..4493
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     protein_bind    5625..5658
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    5709..5742
                     /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)."
     promoter        complement(5761..6939)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          5770..6708
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(5793..5813)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     repeat_region   join(7040..7122,1..47)
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 cgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggaca tgtgagcaaa
       61 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
      121 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
      181 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
      241 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
      301 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
      361 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
      421 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
      481 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
      541 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
      601 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
      661 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
      721 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
      781 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
      841 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
      901 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
      961 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     1021 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     1081 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     1141 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     1201 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     1261 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     1321 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     1381 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     1441 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     1501 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     1561 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     1621 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     1681 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     1741 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     1801 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     1861 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     1921 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     1981 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     2041 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
     2101 gagagtgcac cataaaattg taaacgttaa tattttgtta aaattcgcgt taaatttttg
     2161 ttaaatcagc tcatttttta accaatagac cgaaatcggc aaaatccctt ataaatcaaa
     2221 agaatagccc gagatagagt tgagtgttgt tccagtttgg aacaagagtc cactattaaa
     2281 gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg
     2341 tgaaccatca cccaaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa
     2401 ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa
     2461 ggaagggaag aaagcgaaag gagcgggcgc taaggcgctg gcaagtgtag cggtcacgct
     2521 gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt actatggttg
     2581 ctttgacgta tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
     2641 cgcccctgca ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg
     2701 cgtcgggcga cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg
     2761 gccaactcca tcactagggg ttcctgcggc cgctcggtcc gcacgtgaga tctcatgcct
     2821 gctattgtct tcccaatcct cccccttgct gtcctgcccc accccacccc ccagaataga
     2881 atgacaccta ctcagacaat gcgatgcaat ttcctcattt tattaggaaa ggacagtggg
     2941 agtggcacct tccagggtca aggaaggcac gggggagggg caaacaacag atggctggca
     3001 actagaaggc acagtcgaag atctctcgag cagcgctcgg tatcgatgcg gggaggcggc
     3061 ccaaagggag atccgactcg tctgagggcg aaggcgaaga cgcggaagag gccgcagagc
     3121 cggcagcagg ccgcgggaag gaaggtccgc tggattgagg gccgaaggga cgtagcagaa
     3181 ggacgtcccg cgcagaatcc aggtggcaac ataggcgagc agccaaggaa aggacgatga
     3241 tttccccgac aacaccacgg aattgtcagt gcccaacagc cgagcccctg tccagcagcg
     3301 ggcaaggcag gcggcgatga gttccgccgt ggcaataggg agggggaaag cgaaagtccc
     3361 ggaaaggagc tgacaggtgg tggcaatgcc ccaaccagtg ggggttgcgt cagcaaacac
     3421 agtgcacacc acgccacgtt gcctgacaac gggccacaac tcctcataaa gagacagcaa
     3481 ccaggattta tacaaggagg agaaaatgaa agccatacgg gaagcaatag catgatacaa
     3541 aggcattaaa gcagcgtatc cacatagcgt aaaaggagca acatagttaa gaataccagt
     3601 caatctttca caaattttgt aatccagagg ttgattatcg ataagcttga tatcgaattc
     3661 ataacttcgt atagcataca ttatacgaag ttattctttg caccattcta aagaataaca
     3721 gtgataattt ctgggttaag gcaaataact tcgtatagga tactttatac gaagttatgc
     3781 tagccggatc cgccaccatg gtgagcgagc tgattaagga gaacatgcac atgaagctgt
     3841 acatggaggg caccgtgaac aaccaccact tcaagtgcac atccgagggc gaaggcaagc
     3901 cctacgaggg cacccagacc atgagaatca aggcggtcga gggcggccct ctccccttcg
     3961 ccttcgacat cctggctacc agcttcatgt acggcagcaa aaccttcatc aaccacaccc
     4021 agggcatccc cgacttcttt aagcagtcct tccccgaggg cttcacatgg gagagagtca
     4081 ccacatacga agacgggggc gtgctgaccg ctacccagga caccagcctc caggacggct
     4141 gcctcatcta caacgtcaag atcagagggg tgaacttccc atccaacggc cctgtgatgc
     4201 agaagaaaac actcggctgg gaggcctcca ccgagaccct gtaccccgct gacggcggcc
     4261 tggaaggcag agccgacatg gccctgaagc tcgtgggcgg gggccacctg atctgcaact
     4321 tgaagaccac atacagatcc aagaaacccg ctaagaacct caagatgccc ggcgtctact
     4381 atgtggacag aagactggaa agaatcaagg aggccgacaa agagacctac gtcgagcagc
     4441 acgaggtggc tgtggccaga tactgcgacc tccctagcaa actggggcac agatctggac
     4501 tcagaagtgg aagcggtctc gaggcagctg cgaagaagcg gataacatat atttcctggg
     4561 ccaatgacct cagtcccaag cagatccaac aaattcgaga ggtctctgta aagaacaact
     4621 cctacaatgg tatcacagga gtcctgcttt gtctcaaggg cctgttcttc caaatcttgg
     4681 aaggagacgg cgagcgagta gataagctct acgaaaagat actggttgat gaaaggcatc
     4741 gcgatatact gtgcctgaaa actgagctgg acgtaacgga aaaaatgttc ccagactggg
     4801 gaatgaagac gataaacctg gacgaacagg aggggataat tcttgaacct atcaaaagcc
     4861 tcctctttac catcgctgaa tcacacagaa ttctggagaa atacacccaa ccgtctgtga
     4921 tcaacataat aacgcaaggc attgatccgt tgacggtagc tccccacata gtggaaaaga
     4981 ttattttctt tagcgacatt gtatcctttt ccacatttag tgagaagttg ccggtcgata
     5041 aggtggtaaa cttggtcaac cattatctga caatttgcac ggagataatc agcgcacatg
     5101 gtggagaggt aattaagttc ataggcgatt gtgtaatggc tggattctct acgaaccaag
     5161 cagatgcggc cattagtaca ggtcttgaga tattggctga attgaaaagc cttcgcgagc
     5221 tgacagaagt aaatgaccct cttcatttcc tgtacacggg catcggactc agctgtggta
     5281 atgtcattga aggtaacatt ggatcaccag tcaaaatgga ttacaccctg cttggagacg
     5341 ctgtcaacgt ggcctcacgc ctggaagctc tcagtcggca tttgccctat tcaatgattc
     5401 ttagcaccga agtaaaaaac tactgttgta ccccttggtc tttcattaac ctcgggagac
     5461 accagatgaa gggaaagcag accctggtgg agatttttgc gatcaacgac tccattgtcc
     5521 gcaaatcctc agacgcgagc caaattgcca atttcatcgc gcaggctctc gatatgatcg
     5581 catttaaacc acaaataaag tagactagtt agctagggcg cgccataact tcgtataatg
     5641 tatgctatac gaagttatta agaggtttca tattgctaat agcagctaca atccagctac
     5701 cattctgcat aacttcgtat aaagtatcct atacgaagtt attccggaag atccggtacc
     5761 tcacgacacc tgaaatggaa gaaaaaaact ttgaaccact gtctgaggct tgagaatgaa
     5821 ccaagatcca aactcaaaaa gggcaaattc caaggagaat tacatcaagt gccaagctgg
     5881 cctaacttca gtctccaccc actcagtgtg gggaaactcc atcgcataaa acccctcccc
     5941 ccaacctaaa gacgacgtac tccaaaagct cgagaactaa tcgaggtgcc tggacggcgc
     6001 ccggtactcc gtggagtcac atgaagcgac ggctgaggac ggaaaggccc ttttcctttg
     6061 tgtgggtgac tcacccgccc gctctcccga gcgccgcgtc ctccattttg agctccctgc
     6121 agcagggccg ggaagcggcc atctttccgc tcacgcaact ggtgccgacc gggccagcct
     6181 tgccgcccag ggcggggcga tacacggcgg cgcgaggcca ggcaccagag caggccggcc
     6241 agcttgagac tacccccgtc cgattctcgg tggccgcgct cgcaggcccc gcctcgccga
     6301 acatgtgcgc tgggacgcac gggccccgtc gccgcccgcg gccccaaaaa ccgaaatacc
     6361 agtgtgcaga tcttggcccg catttacaag actatcttgc cagaaaaaaa gcgtcgcagc
     6421 aggtcatcaa aaattttaaa tggctagaga cttatcgaaa gcagcgagac aggcgcgaag
     6481 gtgccaccag attcgcacgc ggcggcccca gcgcccaggc caggcctcaa ctcaagcacg
     6541 aggcgaaggg gctccttaag cgcaaggcct cgaactctcc cacccacttc caacccgaag
     6601 ctcgggatca agaatcacgt actgcagcca ggtggaagta attcaaggca cgcaagggcc
     6661 ataacccgta aagaggccag gcccgcggga accacacacg gcacttacct gtgttctggc
     6721 ggcaaacccg ttgcgaaaaa gaacgttcac ggcgactact gcacttatat acggttctcc
     6781 cccaccctcg ggaaaaaggc ggagccagta cacgacatca ctttcccagt ttaccccgcg
     6841 ccaccttctc taggcaccgg ttcaattgcc gacccctccc cccaacttct cggggactgt
     6901 gggcgatgtg cgctctgccc actgacgggc accggagcca attcccactc ctttcaagac
     6961 ctagaaggtc cattagctgc aaagattcct ctctgtttaa aactttatcc atctttgcaa
     7021 agcttacgcg tgcggccgca ggaaccccta gtgatggagt tggccactcc ctctctgcgc
     7081 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc ga
//
