LOCUS       sequence-325689-        5013 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0459m 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 5013)
  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 5013)
  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: AiP12254-pAAV-AiE0459m-minBG-SYFP2-WPRE3-BGHpA...
FEATURES             Location/Qualifiers
     source          1..5013
                     /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      1025..1034
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1031..1033,1034..1036,1037..1747)
                     /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(1076..1097)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1337..1356)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1684..1705
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1840..1860)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2047..2158
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2276..2416
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2276..2405
                     /label=AAV2 ITR
     rep_origin      2491..2946
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2578..2597)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2788..2809
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2963..2982)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3082..3104
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3142..3160)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3228..3332
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3333..3401,3402..4193)
                     /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(3551..3570)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4364..4952
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4853..4872
                     /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 gcgttgactg tgtcatgact ctcagtgtct cttttttcca
      181 cggttctggg atcatcattt ctttctcttc agacagtcta gatgagcagc cgagtgctta
      241 tctttaaggc tgactctaga cagccagttg ttatttcata aatctgttaa aactcaccgt
      301 gcaaacacca gcgttcctta gctgaccatc ctcaagcaca cgtggagtgt gacctctgct
      361 tccttctctc atttcacata ttagtaatgt gcgtctatga atagttacaa gtgagaaaga
      421 gaaatggaga ttttaccatt taatcatctc cgaagagaag gtgagaagaa attgagaaag
      481 gtaatttgca taccatgcat aggcattaga gaaagcattt tttttcttaa ctctccattt
      541 gtttacagat atttccatat ggccttttaa aaacttcaac agagaaattc gaacacaact
      601 aactggaatt tcaattttca tataaaacca gcatcataga tatatatgta gctactctcc
      661 cacttcacat tattcagtta gtatattttc taagctaatt tccaatccat tttaagacta
      721 tgtacagctc atatgaggcc cccatatgtt cgttcacgtt actttgcatc cgtttacatg
      781 attaaacccc aagaacaaat cgattagaag ccaagagcaa atgcttttgg ctccacatta
      841 gaaaactcaa cagagaaacc tttacaagtg atagtcatct gggtaagaga catgcaaaca
      901 taattaccat tattcgtgga cttaattccc tgagctcggg ctgggcataa aagtcagggc
      961 agagccatct attgcttaca tttgcttctg ggatccagat ctttcgaagc tagcgctacc
     1021 ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     1081 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     1141 tgccacctac ggcaagctga ccctgaagct gatctgcacc accggcaagc tgcccgtgcc
     1201 ctggcccacc ctcgtgacca ccctgggcta cggcgtgcag tgcttcgccc gctaccccga
     1261 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     1321 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     1381 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     1441 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca ccgccgacaa
     1501 gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagg acggcggcgt
     1561 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     1621 cgacaaccac tacctgagct accagtccaa gctgagcaaa gaccccaacg agaagcgcga
     1681 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     1741 gtacaagtaa gtcgacggcg cgccgcggcc gcgaattcga tatcataatc aacctctgga
     1801 ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg
     1861 tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt
     1921 ctcctccttg tataaatcct ggttagttct tgccacggcg gaactcatcg ccgcctgcct
     1981 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg ctcgagagat
     2041 cttcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     2101 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     2161 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     2221 aggattggga agacaatagc aggcatgaga tctcacgtgc ggaccgagcg gccgcaggaa
     2281 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     2341 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     2401 cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     2461 tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg
     2521 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc
     2581 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2641 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2701 ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2761 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2821 actctatctc gggctattct tttgatttat aagggatttt gccgatttcg gtctattggt
     2881 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta
     2941 caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     3001 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     3061 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     3121 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     3181 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     3241 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     3301 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     3361 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     3421 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     3481 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     3541 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     3601 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     3661 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     3721 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     3781 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     3841 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     3901 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     3961 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     4021 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     4081 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     4141 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     4201 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     4261 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     4321 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     4381 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     4441 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     4501 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     4561 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     4621 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     4681 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     4741 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     4801 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     4861 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     4921 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     4981 tcctggcctt ttgctggcct tttgctcaca tgt
//
