LOCUS       sequence-325686-        4985 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0460m is an enhancer sequence designed to drive AAV-mediated 
            transgene expression in L5_ET cortical neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4985)
  AUTHORS   Graybuck LT, Daigle TL, Sedeno-Cortes AE, Walker M, Kalmbach B, Lenz
            GH, Morin E, Nguyen TN, Garren E, Bendrick JL, Kim TK, Zhou T, 
            Mortrud M, Yao S, Siverts LA, Larsen R, Gore BB, Szelenyi ER, Trader
            C, Balaram P, van Velthoven CTJ, Chiang M, Mich JK, Dee N, Goldy J, 
            Cetin AH, Smith K, Way SW, Esposito L, Yao Z, Gradinaru V, Sunkin 
            SM, Lein E, Levi BP, Ting JT, Zeng H, Tasic B
  TITLE     Enhancer viruses for combinatorial cell-subclass-specific labeling.
  JOURNAL   Neuron. 2021 Mar 23. pii: S0896-6273(21)00159-8. doi: 
            10.1016/j.neuron.2021.03.011.
  PUBMED    33789083
REFERENCE   2  (bases 1 to 4985)
  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"; journalName: "Neuron. 
            2021 Mar 23. pii: S0896-6273(21)00159-8. doi: 
            10.1016/j.neuron.2021.03.011."
COMMENT     Sequence Label: AiP12255-pAAV-AiE0460m-minBG-SYFP2-WPRE3-BGHpA...
FEATURES             Location/Qualifiers
     source          1..4985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      997..1006
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1003..1005,1006..1008,1009..1719)
                     /codon_start=1
                     /product="monomeric optimized variant of EYFP, identical to
                     mVenus but with L68V (Kremers et al., 2006)"
                     /label=SYFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1048..1069)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1309..1328)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1656..1677
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1812..1832)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2019..2130
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2248..2388
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2248..2377
                     /label=AAV2 ITR
     rep_origin      2463..2918
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2550..2569)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2760..2781
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2935..2954)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3054..3076
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3114..3132)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3200..3304
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3305..3373,3374..4165)
                     /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(3523..3542)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4336..4924
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4825..4844
                     /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 gcgtttaact tgaggaaatc tgccacaaga tgttctcaag
      181 tgcaaatgtc tcttgatgca tacacgcatg catttctatt taagtatcca acgagaattg
      241 tggggttgag gtaggtattt tagcttataa aaaaaaaata ccttatgagc ttattttgga
      301 tcatataaaa gtatgagcac ttttagaaca cttttaataa tgcaaatcat ctctgcatca
      361 tccaaattta atccatgaac atttagtcat cgttatctta catgagtcat atctgtgtcc
      421 ttattgtctt atcttaaatg cattaattaa ttcgatagag catctataaa aatgaaacgc
      481 tattcaaaag ctggcccatg acaagatcag tgtgttggtt tctatgccta tagtggtgca
      541 atggaaggag tgcgctgtac cccacagact ttgtgattga tgagttgtct actttattca
      601 caatgaagct aaaaacaatc taacaaccat cagggtgcta cagcagcaca gaaatgacta
      661 gctcttttgt gtcaagttat tgccggttgt cttagtcact gtccgactga tgtgcagaga
      721 caccatgact aaggcaacgc ttatcaaaga aagagtctag ttgagagctt gcttgtagtt
      781 ccagaggttt gtttagtcta gcatggtggc acacactgtg ctagaacaat attgagagct
      841 acaacctgat ctgaaaagca gagagagata gagacagaaa gaggcagaga gacactggcc
      901 ctggagctcg ggctgggcat aaaagtcagg gcagagccat ctattgctta catttgcttc
      961 tgggatccag atctttcgaa gctagcgcta ccggtcgcca ccatggtgag caagggcgag
     1021 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
     1081 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
     1141 ctgatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgggc
     1201 tacggcgtgc agtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag
     1261 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
     1321 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     1381 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     1441 aacagccaca acgtctatat caccgccgac aagcagaaga acggcatcaa ggccaacttc
     1501 aagatccgcc acaacatcga ggacggcggc gtgcagctcg ccgaccacta ccagcagaac
     1561 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag ctaccagtcc
     1621 aagctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     1681 gccgccggga tcactctcgg catggacgag ctgtacaagt aagtcgacgg cgcgccgcgg
     1741 ccgcgaattc gatatcataa tcaacctctg gattacaaaa tttgtgaaag attgactggt
     1801 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat
     1861 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttagtt
     1921 cttgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     1981 ttgggcactg acaattccgt ggctcgagag atcttcgact gtgccttcta gttgccagcc
     2041 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     2101 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     2161 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatga
     2221 gatctcacgt gcggaccgag cggccgcagg aacccctagt gatggagttg gccactccct
     2281 ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct
     2341 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc
     2401 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata
     2461 gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     2521 cgctacactt gccagcgcct tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     2581 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     2641 tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg
     2701 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     2761 tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt cttttgattt
     2821 ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt aacaaaaatt
     2881 taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat
     2941 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc
     3001 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     3061 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt
     3121 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     3181 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     3241 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     3301 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     3361 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     3421 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     3481 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     3541 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     3601 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     3661 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     3721 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     3781 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     3841 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     3901 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     3961 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     4021 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     4081 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     4141 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     4201 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     4261 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     4321 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4381 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     4441 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     4501 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     4561 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     4621 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     4681 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     4741 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     4801 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     4861 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     4921 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     4981 catgt
//
