LOCUS       sequence-421885-        6531 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV construct expressing dTomato driven by cholinergic 
            neuron-targeting enhancer. Alias: pAAV_S9E27_dTomato_nlsdTomato.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6531)
  AUTHORS   Furlanis E, Dai M, Leyva Garcia B, Tran T, Vergara J, Pereira A, 
            Gorissen BL, Wills S, Vlachos A, Hairston A, Dwivedi D, Du S, 
            McMahon J, Huang S, Morabito A, Vazquez A, Kim S, Lee AT, Chang EF, 
            Razzaq T, Qazi A, Vargish G, Yuan X, Caccavano A, Hunt S, 
            Chittajallu R, McLean N, Hewitt L, Paranzino E, Rice H, Cummins AC, 
            Plotnikova A, Mohanty A, Tangen AC, Shin JH, Azadi R, Eldridge MAG, 
            Alvarez VA, Averbeck BB, Alyahyay M, Vallejo TR, Soheib M, Vattino 
            LG, MacGregor CP, Chatain CP, Banks E, Olah VJ, Naskar S, Hill S, 
            Liebergall S, Badiani R, Hyde L, Hanley E, Xu Q, Allaway KC, 
            Goldberg EM, Rowan MJM, Nowakowski TJ, Lee S, Favuzzi E, Kaeser PS, 
            Sjulson L, Batista-Brito R, Takesian AE, Ibrahim LA, Iqbal A, Pelkey
            KA, McBain CJ, Dimidschstein J, Fishell G, Wang Y
  TITLE     An enhancer-AAV toolbox to target and manipulate distinct 
            interneuron subtypes.
  JOURNAL   Neuron. 2025 May 21;113(10):1525-1547.e15. doi: 
            10.1016/j.neuron.2025.05.002.
  PUBMED    40403705
REFERENCE   2  (bases 1 to 6531)
  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: 
            "10.1016/j.neuron.2025.05"; journalName: "Neuron"; date: 
            "2025-05-21- 21"; volume: "113"; issue: "10"; pages: "1525-1547.e15"
COMMENT     Sequence Label: pAAV_BiCHATe27_dTomato_nlsdTomato
FEATURES             Location/Qualifiers
     source          1..6531
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     intron          1150..1282
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     regulatory      1305..1314
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1311..2012
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     CDS             2034..2090
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2103..2123
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2124..2822
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGL
                     VTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIHQ
                     ALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRHH
                     LFLYGMDELYK"
     misc_feature    2877..3465
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2930..2950)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3348..3359)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3634..3715
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3671..3690)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3725..3744
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   3794..3934
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3794..3923
                     /label=AAV2 ITR
     rep_origin      4009..4464
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4096..4115)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4306..4327
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4481..4500)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4600..4622
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4660..4678)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4746..4850
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4851..4919,4920..5711)
                     /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(5069..5088)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5882..6470
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6371..6390
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtatcgat cctcgggtaa tttgtaaaga caatttttct
      181 tttccccctc tgaacctagg ccttaggctg gctaggcaag tctccactac agccccgccc
      241 actccatctc ggctgtggct cgcgggaaag gcagccccct acagttagtc tccccagcca
      301 tcttctccgg ttcccctgat gctctcatgg ccccagccat cctagagctg gtgcgccatc
      361 gctcaccttt agcattcggc ctgctggggg tgagcttttg ggttagaagg aacaaggcat
      421 cagatggggt cgtgtttttt tacattccaa tcagttttcg gaggggaaga acagacattt
      481 ttgtcagcta attgcattta attacagaga caatcgctgt ttccgaacgt tgttgtgtcc
      541 actccctcat ccttcaatca gttcaagcac atccggcccc agctgggagc cttttaattg
      601 cataatttgg atttcacttc tggaaagaaa cttgagaaag actccgtgag ggtctccatt
      661 aacacaatca gctccccagg cacccggggc tggccctaat tgacccatct cagctaatgc
      721 ctctttccaa agaacgcccg cgtcatttct ttggcttgtt tgtgctattg tatcctgttt
      781 tagtgtcttg atcttaaaca tctttaaaga gggcactcag tggcttgttt aactggaagt
      841 ggtccagctg gtttggaagg acagtgcatc aggtgatgta agtccaaggg taggaatttc
      901 tggcctctcc ttcccctgcc cattgcaccg ctctccccaa caggaagtag ctacacatga
      961 taaggtgcac tgtggcttct aggacttatc gatgtttaaa cttcctgcag cccgggctgg
     1021 gcataaaagt cagggcagag ccatctattg cttacatttg cttctagcct gcaagtcgag
     1081 gagcgcagcc ttccagaagc agagcgcggc gccttaagct gcagaagttg gtcgtgaggc
     1141 actgggcacg taagtatcaa ggttacaaga caggtttaag gagaccaata gaaactgggc
     1201 ttgtcgagac agagaagact cttgcgtttc tgataggcac ctattggtct tactgacatc
     1261 cactttgcct ttctctccac agcgagctcg ctagcgcgat cgctgccacc atggtgagca
     1321 agggcgagga ggtcatcaaa gagttcatgc gcttcaaggt gcgcatggag ggctccatga
     1381 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
     1441 ccgccaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc
     1501 cccagttcat gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc cccgattaca
     1561 agaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     1621 gtctggtgac cgtgacccag gactcctccc tgcaggacgg cacgctgatc tacaaggtga
     1681 agatgcgcgg caccaacttc ccccccgacg gccccgtaat gcagaagaag accatgggct
     1741 gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc gagatccacc
     1801 aggccctgaa gctgaaggac ggcggccact acctggtgga gttcaagacc atctacatgg
     1861 ccaagaagcc cgtgcaactg cccggctact actacgtgga caccaagctg gacatcacct
     1921 cccacaacga ggactacacc atcgtggaac agtacgagcg ctccgagggc cgccaccacc
     1981 tgttcctgta cggcatggac gagctgtaca agtccggaga taacggaagc ggagccacta
     2041 acttctccct gttgaaacaa gcaggggatg tcgaagagaa tcccgggcca tctagaatgg
     2101 cccccaagaa gaagaggaag gtggtgagca agggcgagga ggtcatcaaa gagttcatgc
     2161 gcttcaaggt gcgcatggag ggctccatga acggccacga gttcgagatc gagggcgagg
     2221 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggcggcc
     2281 ccctgccctt cgcctgggac atcctgtccc cccagttcat gtacggctcc aaggcgtacg
     2341 tgaagcaccc cgccgacatc cccgattaca agaagctgtc cttccccgag ggcttcaagt
     2401 gggagcgcgt gatgaacttc gaggacggcg gtctggtgac cgtgacccag gactcctccc
     2461 tgcaggacgg cacgctgatc tacaaggtga agatgcgcgg caccaacttc ccccccgacg
     2521 gccccgtaat gcagaagaag accatgggct gggaggcctc caccgagcgc ctgtaccccc
     2581 gcgacggcgt gctgaagggc gagatccacc aggccctgaa gctgaaggac ggcggccact
     2641 acctggtgga gttcaagacc atctacatgg ccaagaagcc cgtgcaactg cccggctact
     2701 actacgtgga caccaagctg gacatcacct cccacaacga ggactacacc atcgtggaac
     2761 agtacgagcg ctccgagggc cgccaccacc tgttcctgta cggcatggac gagctgtaca
     2821 agtaagtcga cggcgcgccc cacccctgca gggaattcga tatcaagctt atcgataatc
     2881 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     2941 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     3001 ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc
     3061 ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt
     3121 ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg
     3181 ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg
     3241 gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct
     3301 atgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc
     3361 cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc
     3421 ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcatcga taccgagcgc
     3481 tgctcgagag atctgtgata gcggccatca agctggccgc gactctagat cataatcagc
     3541 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
     3601 ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt
     3661 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     3721 agttgtggtt tgtccaaact catcaatgta tcagcttatc gataccgcat gcacgtgcgg
     3781 accgagcggc cgcaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc
     3841 tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc
     3901 tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc tgatgcggta ttttctcctt
     3961 acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta
     4021 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     4081 gcgccttagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     4141 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     4201 acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat
     4261 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     4321 aaactggaac aacactcaac tctatctcgg gctattcttt tgatttataa gggattttgc
     4381 cgatttcggt ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     4441 acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg
     4501 catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     4561 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     4621 ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt
     4681 tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa
     4741 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     4801 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     4861 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     4921 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     4981 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     5041 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     5101 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     5161 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     5221 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     5281 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     5341 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     5401 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     5461 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     5521 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     5581 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     5641 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     5701 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     5761 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     5821 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     5881 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     5941 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     6001 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     6061 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     6121 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     6181 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     6241 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     6301 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     6361 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     6421 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     6481 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg t
//
