LOCUS       sequence-333371-        7186 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV plasmid containing a mKate2 and a truncated PDE4D3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7186)
  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 7186)
  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-PDE4D3-Cat
FEATURES             Location/Qualifiers
     source          1..7186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(20..39)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(699..1490,1491..1559))
                     /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     1322..1341
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1560..1664)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1732..1750
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1788..1810)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1910..1929
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(1946..2401)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2083..2104)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2295..2314
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2476..2616
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2487..2616
                     /label=AAV2 ITR
     polyA_signal    2734..2845
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     misc_feature    2879..3467
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             2985..2996
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     3394..3414
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(3492..3525)
                     /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(3576..3609)
                     /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      3623..3632
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3629..4324
                     /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    5520..5553
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    5604..5637
                     /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(5656..6834)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          5665..6603
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(5688..5708)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     repeat_region   6935..7064
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(join(7126..7186,1..528))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
       61 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
      121 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
      181 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
      241 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
      301 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
      361 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
      421 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
      481 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
      541 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
      601 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
      661 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
      721 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
      781 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
      841 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
      901 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
      961 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     1021 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     1081 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     1141 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     1201 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     1261 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     1321 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     1381 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     1441 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     1501 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     1561 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     1621 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     1681 gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt
     1741 atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg
     1801 cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt
     1861 cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag
     1921 cagattgtac tgagagtgca ccataaaatt gtaaacgtta atattttgtt aaaattcgcg
     1981 ttaaattttt gttaaatcag ctcatttttt aaccaataga ccgaaatcgg caaaatccct
     2041 tataaatcaa aagaatagcc cgagatagag ttgagtgttg ttccagtttg gaacaagagt
     2101 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     2161 ggcccactac gtgaaccatc acccaaatca agttttttgg ggtcgaggtg ccgtaaagca
     2221 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     2281 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctaaggcgct ggcaagtgta
     2341 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     2401 tactatggtt gctttgacgt atgcggtgtg aaataccgca cagatgcgta aggagaaaat
     2461 accgcatcag gcgcccctgc aggcagctgc gcgctcgctc gctcactgag gccgcccggg
     2521 caaagcccgg gcgtcgggcg acctttggtc gcccggcctc agtgagcgag cgagcgcgca
     2581 gagagggagt ggccaactcc atcactaggg gttcctgcgg ccgctcggtc cgcacgtgag
     2641 atctcatgcc tgctattgtc ttcccaatcc tcccccttgc tgtcctgccc caccccaccc
     2701 cccagaatag aatgacacct actcagacaa tgcgatgcaa tttcctcatt ttattaggaa
     2761 aggacagtgg gagtggcacc ttccagggtc aaggaaggca cgggggaggg gcaaacaaca
     2821 gatggctggc aactagaagg cacagtcgaa gatctctcga gcagcgctcg gtatcgatgc
     2881 ggggaggcgg cccaaaggga gatccgactc gtctgagggc gaaggcgaag acgcggaaga
     2941 ggccgcagag ccggcagcag gccgcgggaa ggaaggtccg ctggattgag ggccgaaggg
     3001 acgtagcaga aggacgtccc gcgcagaatc caggtggcaa cataggcgag cagccaagga
     3061 aaggacgatg atttccccga caacaccacg gaattgtcag tgcccaacag ccgagcccct
     3121 gtccagcagc gggcaaggca ggcggcgatg agttccgccg tggcaatagg gagggggaaa
     3181 gcgaaagtcc cggaaaggag ctgacaggtg gtggcaatgc cccaaccagt gggggttgcg
     3241 tcagcaaaca cagtgcacac cacgccacgt tgcctgacaa cgggccacaa ctcctcataa
     3301 agagacagca accaggattt atacaaggag gagaaaatga aagccatacg ggaagcaata
     3361 gcatgataca aaggcattaa agcagcgtat ccacatagcg taaaaggagc aacatagtta
     3421 agaataccag tcaatctttc acaaattttg taatccagag gttgattatc gataagcttg
     3481 atatcgaatt cataacttcg tatagcatac attatacgaa gttattcttt gcaccattct
     3541 aaagaataac agtgataatt tctgggttaa ggcaaataac ttcgtatagg atactttata
     3601 cgaagttatg ctagccggat ccgccaccat ggtgagcgag ctgattaagg agaacatgca
     3661 catgaagctg tacatggagg gcaccgtgaa caaccaccac ttcaagtgca catccgaggg
     3721 cgaaggcaag ccctacgagg gcacccagac catgagaatc aaggcggtcg agggcggccc
     3781 tctccccttc gccttcgaca tcctggctac cagcttcatg tacggcagca aaaccttcat
     3841 caaccacacc cagggcatcc ccgacttctt taagcagtcc ttccccgagg gcttcacatg
     3901 ggagagagtc accacatacg aagacggggg cgtgctgacc gctacccagg acaccagcct
     3961 ccaggacggc tgcctcatct acaacgtcaa gatcagaggg gtgaacttcc catccaacgg
     4021 ccctgtgatg cagaagaaaa cactcggctg ggaggcctcc accgagaccc tgtaccccgc
     4081 tgacggcggc ctggaaggca gagccgacat ggccctgaag ctcgtgggcg ggggccacct
     4141 gatctgcaac ttgaagacca catacagatc caagaaaccc gctaagaacc tcaagatgcc
     4201 cggcgtctac tatgtggaca gaagactgga aagaatcaag gaggccgaca aagagaccta
     4261 cgtcgagcag cacgaggtgg ctgtggccag atactgcgac ctccctagca aactggggca
     4321 cagatctgga ctcagaagtg gaagcggtct cgagatgtct cagatcagtg gagtcaagaa
     4381 attgatgcac agctctagtc tgactaattc aagtatccca aggtttggag ttaaaactga
     4441 acaagaagat gtccttgcca aggaactaga agatgtgaac aaatggggtc ttcatgtttt
     4501 cagaatagca gagttgtctg gtaaccggcc cttgactgtt atcatgcaca ccatttttca
     4561 ggaacgggat ttattaaaaa catttaaaat tccagtagat actttaatta catatcttat
     4621 gactctcgaa gaccattacc atgctgatgt ggcctatcac aacaatatcc atgctgcaga
     4681 tgttgtccag tctactcatg tgctattatc tacacctgct ttggaggctg tgtttacaga
     4741 tttggagatt cttgcagcaa tttttgccag tgcaatacat gatgtagatc atcctggtgt
     4801 gtccaatcaa tttctgatca atacaaactc tgaacttgcc ttgatgtaca atgattcctc
     4861 agtcttagag aaccatcatt tggctgtggg ctttaaattg cttcaggaag aaaactgtga
     4921 cattttccag aatttgacca aaaaacaaag acaatcttta aggaaaatgg tcattgacat
     4981 cgtacttgca acagatatgt caaaacacat gaatctactg gctgatttga agactatggt
     5041 tgaaactaag aaagtgacaa gctctggagt tcttcttctt gataattatt ccgataggat
     5101 tcaggttctt cagaatatgg tgcactgtgc agatctgagc aacccaacaa agcctctcca
     5161 gctgtaccgc cagtggacgg accggataat ggaggagttc ttccgccaag gagaccgaga
     5221 gagggaacgt ggcatggaga taagccccat gtgtgacaag cacaatgctt ccgtggaaaa
     5281 atcacaggtg ggcttcatag actatattgt tcatcccctc tgggagacat gggcagacct
     5341 cgtccaccct gacgcccagg atattttgga cactttggag gacaatcgtg aatggtacca
     5401 gagcacaatc cctcagagcc cctctcctgc acctgatgac ccagaggagg gccggcaggg
     5461 tcaaactgag aaattccagt ttgaactaac ttaagcttac tagttagcta gggcgcgcca
     5521 taacttcgta taatgtatgc tatacgaagt tattaagagg tttcatattg ctaatagcag
     5581 ctacaatcca gctaccattc tgcataactt cgtataaagt atcctatacg aagttattcc
     5641 ggaagatccg gtacctcacg acacctgaaa tggaagaaaa aaactttgaa ccactgtctg
     5701 aggcttgaga atgaaccaag atccaaactc aaaaagggca aattccaagg agaattacat
     5761 caagtgccaa gctggcctaa cttcagtctc cacccactca gtgtggggaa actccatcgc
     5821 ataaaacccc tccccccaac ctaaagacga cgtactccaa aagctcgaga actaatcgag
     5881 gtgcctggac ggcgcccggt actccgtgga gtcacatgaa gcgacggctg aggacggaaa
     5941 ggcccttttc ctttgtgtgg gtgactcacc cgcccgctct cccgagcgcc gcgtcctcca
     6001 ttttgagctc cctgcagcag ggccgggaag cggccatctt tccgctcacg caactggtgc
     6061 cgaccgggcc agccttgccg cccagggcgg ggcgatacac ggcggcgcga ggccaggcac
     6121 cagagcaggc cggccagctt gagactaccc ccgtccgatt ctcggtggcc gcgctcgcag
     6181 gccccgcctc gccgaacatg tgcgctggga cgcacgggcc ccgtcgccgc ccgcggcccc
     6241 aaaaaccgaa ataccagtgt gcagatcttg gcccgcattt acaagactat cttgccagaa
     6301 aaaaagcgtc gcagcaggtc atcaaaaatt ttaaatggct agagacttat cgaaagcagc
     6361 gagacaggcg cgaaggtgcc accagattcg cacgcggcgg ccccagcgcc caggccaggc
     6421 ctcaactcaa gcacgaggcg aaggggctcc ttaagcgcaa ggcctcgaac tctcccaccc
     6481 acttccaacc cgaagctcgg gatcaagaat cacgtactgc agccaggtgg aagtaattca
     6541 aggcacgcaa gggccataac ccgtaaagag gccaggcccg cgggaaccac acacggcact
     6601 tacctgtgtt ctggcggcaa acccgttgcg aaaaagaacg ttcacggcga ctactgcact
     6661 tatatacggt tctcccccac cctcgggaaa aaggcggagc cagtacacga catcactttc
     6721 ccagtttacc ccgcgccacc ttctctaggc accggttcaa ttgccgaccc ctccccccaa
     6781 cttctcgggg actgtgggcg atgtgcgctc tgcccactga cgggcaccgg agccaattcc
     6841 cactcctttc aagacctaga aggtccatta gctgcaaaga ttcctctctg tttaaaactt
     6901 tatccatctt tgcaaagctt acgcgtgcgg ccgcaggaac ccctagtgat ggagttggcc
     6961 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     7021 ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggacatgt gagcaaaagg
     7081 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     7141 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtgg
//
