LOCUS       sequence-325687-        4631 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0462m 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 4631)
  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 4631)
  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: AiP12256-pAAV-AiE0462m-minBG-SYFP2-WPRE3-BGHpA...
FEATURES             Location/Qualifiers
     source          1..4631
                     /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      643..652
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(649..651,652..654,655..1365)
                     /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(694..715)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(955..974)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1302..1323
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1458..1478)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    1665..1776
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   1894..2034
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1894..2023
                     /label=AAV2 ITR
     rep_origin      2109..2564
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2196..2215)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2406..2427
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2581..2600)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2700..2722
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2760..2778)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2846..2950
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2951..3019,3020..3811)
                     /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(3169..3188)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3982..4570
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4471..4490
                     /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 gcgtgctggt gggtgggaag agtttagtaa ctgctcacca
      181 atggtgctgt agacattatc agttgcaaga agtgttttaa tttcagagaa attaaaatat
      241 gcaaaatatg tgtctctgaa tcaacaaaac atggtgcctc aggtttccgt atctgtgtgg
      301 gcctgttttt aggttttctg ttctatctat tcctgggctt ccaccacact gtgttaatga
      361 ggataacttt aaaatacatc ttgatattta gtagagcgca tttccacgcc tgattcatca
      421 cgagcacctt aacttgtctt ggttcttctg cggagatgtt ggcaccagct tcttcacccg
      481 tgactccagc ttctggggaa cctgtgctct cacccctgcc ccgcctgtgc ctgccaggca
      541 caccttaatg agctcgggct gggcataaaa gtcagggcag agccatctat tgcttacatt
      601 tgcttctggg atccagatct ttcgaagcta gcgctaccgg tcgccaccat ggtgagcaag
      661 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
      721 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
      781 ctgaagctga tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
      841 ctgggctacg gcgtgcagtg cttcgcccgc taccccgacc acatgaagca gcacgacttc
      901 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
      961 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     1021 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     1081 aactacaaca gccacaacgt ctatatcacc gccgacaagc agaagaacgg catcaaggcc
     1141 aacttcaaga tccgccacaa catcgaggac ggcggcgtgc agctcgccga ccactaccag
     1201 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagctac
     1261 cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     1321 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaagt cgacggcgcg
     1381 ccgcggccgc gaattcgata tcataatcaa cctctggatt acaaaatttg tgaaagattg
     1441 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     1501 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     1561 ttagttcttg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     1621 cggctgttgg gcactgacaa ttccgtggct cgagagatct tcgactgtgc cttctagttg
     1681 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     1741 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     1801 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     1861 gcatgagatc tcacgtgcgg accgagcggc cgcaggaacc cctagtgatg gagttggcca
     1921 ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc
     1981 cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc
     2041 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca
     2101 accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     2161 cgtgaccgct acacttgcca gcgccttagc gcccgctcct ttcgctttct tcccttcctt
     2221 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     2281 ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg
     2341 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     2401 taatagtgga ctcttgttcc aaactggaac aacactcaac tctatctcgg gctattcttt
     2461 tgatttataa gggattttgc cgatttcggt ctattggtta aaaaatgagc tgatttaaca
     2521 aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag
     2581 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     2641 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     2701 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     2761 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc
     2821 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     2881 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     2941 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     3001 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     3061 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     3121 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     3181 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     3241 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     3301 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     3361 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     3421 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     3481 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     3541 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     3601 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     3661 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     3721 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     3781 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     3841 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     3901 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     3961 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     4021 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     4081 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     4141 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     4201 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     4261 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     4321 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     4381 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     4441 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     4501 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     4561 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     4621 tgctcacatg t
//
