LOCUS       sequence-462733-        4655 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0458m_3xC2 is an enhancer sequence, designed to drive 
            AAV-mediated transgene expression in cortical L5_ET neurons 
            (strong).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4655)
  AUTHORS   Ben-Simon Y, Hooper M, Narayan S, Daigle TL, Dwivedi D, Way SW, 
            Oster A, Stafford DA, Mich JK, Taormina MJ, Martinez RA, Opitz-Araya
            X, Roth JR, Alexander JR, Allen S, Amster A, Arbuckle J, Ayala A, 
            Baker PM, Bakken TE, Barcelli T, Barta S, Bendrick J, Bertagnolli D,
            Bielstein C, Bishwakarma P, Bowlus J, Boyer G, Brouner K, Casian B, 
            Casper T, Chakka AB, Chakrabarty R, Chance RK, Chavan S, Clark M, 
            Colbert K, Collman F, Daniel S, Departee M, DiValentin P, Donadio N,
            Dotson N, Egdorf T, Fliss T, Gabitto M, Garcia J, Gary A, Gasperini 
            M, Gloe J, Goldy J, Gore BB, Graybuck L, Greisman N, Haeseleer F, 
            Halterman C, Haradon Z, Hastings SD, Helback O, Ho W, Hockemeyer D, 
            Huang C, Huff S, Hunker A, Johansen N, Jones D, Juneau Z, Kalmbach 
            B, Kannan M, Khem S, Kussick E, Kutsal R, Larsen R, Lee C, Lee AY, 
            Leibly M, Lenz GH, Li S, Liang E, Lusk N, Madigan Z, Malloy J, 
            Malone J, McCue R, Melchor J, Mollenkopf T, Moosman S, Morin E, 
            Newman D, Ng L, Ngo K, Omstead V, Otto S, Oyama A, Pena N, Pham T, 
            Phillips E, Pom CA, Potekhina L, Ransford S, Ray PL, Rette D, 
            Reynoldson C, Rimorin C, Rocha D, Ruiz A, Sanchez REA, Sawyer L, 
            Sedeno-Cortes A, Sevigny JP, Shapovalova N, Shepard N, Shulga L, 
            Sigler AR, Siverts L, Soliman S, Somasundaram S, Staats B, Stewart 
            K, Szelenyi E, Tieu M, Trader C, Tran A, van Velthoven CTJ, Walker 
            M, Wang Y, Weed N, Wirthlin M, Wood T, Wynalda B, Yao Z, Zhou T, 
            Ariza J, Dee N, Reding M, Ronellenfitch K, Mufti S, Sunkin SM, Smith
            KA, Esposito L, Waters J, Thyagarajan B, Yao S, Lein ES, Zeng H, 
            Levi BP, Ngai J, Ting JT, Tasic B
  TITLE     A suite of enhancer AAVs and transgenic mouse lines for genetic 
            access to cortical cell types.
  JOURNAL   Cell. 2025 May 16:S0092-8674(25)00513-6. doi: 
            10.1016/j.cell.2025.05.002.
  PUBMED    40403729
REFERENCE   2  (bases 1 to 4655)
  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: "Cell. 2025 
            May 16:S0092-8674(25)00513-6. doi: 10.1016/j.cell.2025.05.002."
COMMENT     Sequence Label: AiP13900 -
            pAAV-AiE0458m_3xC2-minBG-SYFP2-WPRE...
FEATURES             Location/Qualifiers
     source          1..4655
                     /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      667..676
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(673..675,676..678,679..1389)
                     /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(718..739)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(979..998)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1326..1347
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1482..1502)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    1689..1800
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   1918..2058
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1918..2047
                     /label=AAV2 ITR
     rep_origin      2133..2588
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2220..2239)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2430..2451
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2605..2624)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2724..2746
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2784..2802)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2870..2974
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2975..3043,3044..3835)
                     /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(3193..3212)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4006..4594
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4495..4514
                     /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 gcgtaatgga aatttccacg gataaataat tcatgagact
      181 tgaggaaaat gtgtgctagt cctcaatacc ttataaaatg tattattcat tttgaattag
      241 attaaaatgt ttaaaaaatg ctatctataa gacactgtgg gagccaaaat ggaaatttcc
      301 acggataaat aattcatgag acttgaggaa aatgtgtgct agtcctcaat accttataaa
      361 atgtattatt cattttgaat tagattaaaa tgtttaaaaa atgctatcta taagacactg
      421 tgggagccaa aatggaaatt tccacggata aataattcat gagacttgag gaaaatgtgt
      481 gctagtcctc aataccttat aaaatgtatt attcattttg aattagatta aaatgtttaa
      541 aaaatgctat ctataagaca ctgtgggagc caagagctcg ggctgggcat aaaagtcagg
      601 gcagagccat ctattgctta catttgcttc tgggatccag atctttcgaa gctagcgcta
      661 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      721 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      781 gatgccacct acggcaagct gaccctgaag ctgatctgca ccaccggcaa gctgcccgtg
      841 ccctggccca ccctcgtgac caccctgggc tacggcgtgc agtgcttcgc ccgctacccc
      901 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      961 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     1021 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1081 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat caccgccgac
     1141 aagcagaaga acggcatcaa ggccaacttc aagatccgcc acaacatcga ggacggcggc
     1201 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1261 cccgacaacc actacctgag ctaccagtcc aagctgagca aagaccccaa cgagaagcgc
     1321 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1381 ctgtacaagt aagtcgacgg cgcgccgcgg ccgcgaattc gatatcataa tcaacctctg
     1441 gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta
     1501 tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt
     1561 ttctcctcct tgtataaatc ctggttagtt cttgccacgg cggaactcat cgccgcctgc
     1621 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggctcgagag
     1681 atcttcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     1741 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     1801 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     1861 ggaggattgg gaagacaata gcaggcatga gatctcacgt gcggaccgag cggccgcagg
     1921 aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg
     1981 ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag
     2041 cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     2101 tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc
     2161 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc
     2221 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     2281 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     2341 acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     2401 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     2461 caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg
     2521 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt
     2581 tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     2641 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     2701 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     2761 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     2821 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     2881 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     2941 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     3001 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     3061 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     3121 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     3181 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     3241 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     3301 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     3361 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     3421 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     3481 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     3541 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     3601 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     3661 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     3721 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     3781 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     3841 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     3901 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     3961 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     4021 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     4081 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     4141 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     4201 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     4261 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     4321 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     4381 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     4441 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     4501 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     4561 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     4621 gttcctggcc ttttgctggc cttttgctca catgt
//
