LOCUS       sequence-384897-        7417 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV plasmid for mammalian Cre-dependent expression of the T391A 
            mutant of tAKAR-alpha (abolished sensor phosphorylation site)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7417)
  AUTHORS   Ma L, Day-Cooney J, Benavides OJ, Muniak MA, Qin M, Ding JB, Mao T, 
            Zhong H
  TITLE     Locomotion activates PKA through dopamine and adenosine in striatal 
            neurons.
  JOURNAL   Nature. 2022 Nov;611(7937):762-768. doi: 10.1038/s41586-022-05407-4.
            Epub 2022 Nov 9.
  PUBMED    36352228
REFERENCE   2  (bases 1 to 7417)
  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/s41586-022-05407-4"; journalName: "Nature"; date: 
            "2022-11"; volume: "611"; issue: "7937"; pages: "762-768"
COMMENT     Sequence Label: pAAV_SynP_DIO_tAKAR-alpha- T391A_WPRE
FEATURES             Location/Qualifiers
     source          1..7417
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   7..136
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     protein_bind    complement(628..661)
                     /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)."
     protein_bind    complement(712..745)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(1393..1905,1906..1908,1909..1911))
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNWNSVNVYIMADKQKNGIK
                     VNFKIRHNIE"
     CDS             complement(join(2815..3327,3328..3330,3331..3333))
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     regulatory      3330..3339
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3408..3441
                     /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)."
     protein_bind    3492..3525
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3550..4138
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3603..3623)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4021..4032)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4170..4646
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4369..4388)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4686..4826
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4686..4815
                     /label=AAV2 ITR
     rep_origin      4901..5356
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4988..5007)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5198..5219
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5373..5392)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5492..5514
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5552..5570)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5638..5742
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5743..5811,5812..6603)
                     /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(5961..5980)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6774..7362
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7263..7282
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acatgtcctg caggcagctg cgcgctcgct cgctcactga ggccgcccgg gcgtcgggcg
       61 acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc
      121 atcactaggg gttcctgcgg ccgcacgcgt taactgcaga gggccctgcg tatgagtgca
      181 agtgggtttt aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga
      241 cccactggac aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg
      301 gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt
      361 gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac
      421 gtcactcgcc ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc
      481 cgccggccca gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca
      541 tctgcgctgc ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt
      601 cgtgtcgtgc ctgagagcgc gtccggaata acttcgtata ggatacttta tacgaagtta
      661 tgcagaatgg tagctggatt gtagctgcta ttagcaatat gaaacctctt aataacttcg
      721 tatagcatac attatacgaa gttatggcgc gccaagctta ccggtggatc cttacaagcc
      781 ctgcttagcg agtgcagcag tgacatcctc agccaaagtg gcaagctgag gagattcgag
      841 caggttgatg cttccagacg aggagacatt gctgagtcgt cggggtgatg ccacgctgga
      901 tctgcctggg gactgtgtaa tgatctcagc cccatggtcc acacgggctt tagcatgctc
      961 tctgaagttc aacttttggc tgtcaatctt gacattacca cctccaggta catgatgagc
     1021 attatcaaga gaaccaactt tagcttgggc cttttctttg aaatctagtt ttacactctc
     1081 aattttcaca cgtccgccac ctggcctgtg gcggatgttc ttcagagagc cacatttgga
     1141 tgtcacatgg cttaggtcta ttttcttggt aacaatttgt acctgccccc ctttaggctg
     1201 gtatttgatg ttgtctgttg atccgatttt ggatttgaca ttcttcaggt ctggcagtgg
     1261 ttggttaata agccgaagct gcttgggagt cgcaggagat tttggaggag tacgtatgat
     1321 ggcgaccttc ttctcactcg gcaccaagat ggcagacttg ggggttcctg gggtaccctt
     1381 gtacactccg gactcgatgt tgtggcggat cttgaagttc accttgatgc cgttcttctg
     1441 cttgtcggcc atgatataga cgttgacgct gttccagttg tactccagct tgtgccccag
     1501 gatgttgccg tcctccttga agtcgatgcc cttcagctcg atgcggttca ccagggtgtc
     1561 gccctcgaac ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga agaagatggt
     1621 gcgctcctgg acgtagcctt cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg
     1681 gtcggggtag cgggcgaagc acatcaggcc gtagccgaag gtggtcacga gggtgggcca
     1741 gggcacgggc agcttgccgg tggtgcagat gagcttcagg gtcagcttgc cgtaggtggc
     1801 atcgccctcg ccctcgccgg acacgctgaa cttgtggccg tttacgtcgc cgtccagctc
     1861 gaccaggatg ggcaccaccc cggtgaacag ctcctcgccc ttgctcacca ttgaacctcc
     1921 agttccaccc ttgtatagct cgtccatgcc gagagtgatc ccggcggcgg tcactctctc
     1981 cagcaggacc atgtgatcgc gcttctcgtt ggggtctttg ctcagggcgg actggtagct
     2041 caggtagtgg ttgtcgggca gcagcacggg gccgtcgccg atgggggtgt tctgctggta
     2101 gtggtcggcg agctgcacgc tgcccatgag ctcgctgccg ccggtgccgc cgtccaccag
     2161 ggcggcgcga cgcaggccct tggtgctgcc ctcgccgctg cctggcttac ctgcagagcg
     2221 atcaactttg ttctgctcga ggcactgctt aaatttgtcg tttatgaaaa tgaccagaga
     2281 taaaatatct gattccacgc ctacaccaac ggttatttca tcaccttgag acagtaactg
     2341 attgctgttc ttctcgactt tttgcccatt taaccaggtc ccattagtgg aaatgtcatt
     2401 caataaaagg ttaccgtctt ctcctagtag tatttggaaa tgcttatttg acagtcttga
     2461 aatgtttcct aaatgatagt cacaggctgg gtttctacca aatgtccaaa ctttctttat
     2521 ggatcgtttt tccttaagca cttgtgaaat atcagctgac aaatctcgga tgggaatttg
     2581 acccgtggta caaatgaccc tgcacacaat gttttcgccg atctgttctt gagaaaactt
     2641 atgcatgcgg gcggcggtca cgaactccag caggaccatg tgatcgcgct tctcgttggg
     2701 gtctttgctc agcttggact gggtgctcag gtagtggttg tcgggcagca gcacggggcc
     2761 gtcgccgatg ggggtgttct gctggtagtg gtcggcgagc tgcacgctgc cgtcctcgat
     2821 gttgtggcgg atcttgaagt tcaccttgat gccgttcttc tgcttgtcgg ccatgatata
     2881 gacgttgtgg ctgttgtagt tgtactccag cttgtgcccc aggatgttgc cgtcctcctt
     2941 gaagtcgatg cccttcagct cgatgcggtt caccagggtg tcgccctcga acttcacctc
     3001 ggcgcgggtc ttgtagttgc cgtcgtcctt gaagaagatg gtgcgctcct ggacgtagcc
     3061 ttcgggcatg gcggacttga agaagtcgtg ctgcttcatg tggtcggggt agcggctgaa
     3121 gcactgcacg ccgtaggtca gggtggtcac gagggtgggc cagggcacgg gcagcttgcc
     3181 ggtggtgcag atgaacttca gggtcagctt gccgtaggtg gcatcgccct cgccctcgcc
     3241 ggacacgctg aacttgtggc cgtttacgtc gccgtccagc tcgaccagga tgggcaccac
     3301 cccggtgaac agctcctcgc ccttgctcac catggtggcg accggtctcg agcccgggaa
     3361 ttcaatcgat agaaccgagg tgcggtggct ctagagtcga cgctagcata acttcgtata
     3421 aagtatccta tacgaagtta tttgccttaa cccagaaatt atcactgtta ttctttagaa
     3481 tggtgcaaag aataacttcg tataatgtat gctatacgaa gttatgaatt cgatatcaag
     3541 cttatcgata atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     3601 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     3661 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat
     3721 gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca
     3781 acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc
     3841 cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg
     3901 gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct
     3961 tggctgctcg cctatgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct
     4021 tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt
     4081 ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat
     4141 cgataccgag cgctgctcga gagatctacg ggtggcatcc ctgtgacccc tccccagtgc
     4201 ctctcctggc cctggaagtt gccactccag tgcccaccag ccttgtccta ataaaattaa
     4261 gttgcatcat tttgtctgac taggtgtcct tctataatat tatggggtgg aggggggtgg
     4321 tatggagcaa ggggcaagtt gggaagacaa cctgtagggc ctgcggggtc tattgggaac
     4381 caagctggag tgcagtggca caatcttggc tcactgcaat ctccgcctcc tgggttcaag
     4441 cgattctcct gcctcagcct cccgagttgt tgggattcca ggcatgcatg accaggctca
     4501 gctaattttt gtttttttgg tagagacggg gtttcaccat attggccagg ctggtctcca
     4561 actcctaatc tcaggtgatc tacccacctt ggcctcccaa attgctggga ttacaggcgt
     4621 gaaccactgc tcccttccct gtccttctga ttttgtaggt aaccacgtgc ggaccgagcg
     4681 gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac
     4741 tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag
     4801 cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct
     4861 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
     4921 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta
     4981 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     5041 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     5101 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     5161 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     5221 acaacactca actctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     5281 gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     5341 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     5401 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     5461 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     5521 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     5581 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     5641 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     5701 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     5761 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     5821 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     5881 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     5941 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     6001 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     6061 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     6121 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     6181 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     6241 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     6301 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     6361 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     6421 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     6481 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     6541 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     6601 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     6661 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     6721 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     6781 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     6841 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     6901 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     6961 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     7021 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     7081 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     7141 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     7201 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     7261 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     7321 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     7381 tttttacggt tcctggcctt ttgctggcct tttgctc
//
