LOCUS       sequence-473584-        5214 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0976h is an enhancer sequence, designed to drive AAV-mediated 
            transgene expression in specific populations of brain cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5214)
  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 5214)
  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: AiP13460 -
            pAAV-AiE0976h-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5214
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     CDS             complement(217..228)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      680..689
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(686..688,689..691,692..1402)
                     /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(992..1011)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1339..1360
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1412..1468
                     /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             1481..1546
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1577..1954
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(2041..2061)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2248..2359
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2477..2617
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2477..2606
                     /label=AAV2 ITR
     rep_origin      2692..3147
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2779..2798)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2989..3010
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3164..3183)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3283..3305
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3343..3361)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3429..3533
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3534..3602,3603..4394)
                     /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(3752..3771)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4565..5153
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5054..5073
                     /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 gcgtggttca ctggatttga atggcgtttg agaaggggag
      181 aaggtggcgc aagtgaacca aagcaactta actccatctg ccttcgattc caaggctgag
      241 aaaacatttc cagaaggcct ggtgcataag aacctgatgt ggtgacagga tgttctgtct
      301 tcctcttgtt acacaagcaa gcgcgctgag tctcggcagg ctcataaatc ctgagggcct
      361 tttccagcag ccccgttcgg gtcgcctacc cccattccag gaaagggacg ctttgaaacc
      421 ctcatcacaa gagaacattg tgtctttgaa catccggcgc cactggggtt tgacttctcc
      481 cagcgcaggt aggatgggcc agccagtgag gtcagatgtg atttatgctt ctaaagctct
      541 tgcctttcag aattttggaa gaaaatacaa gctgcccaat actgcagagc tcgggctggg
      601 cataaaagtc agggcagagc catctattgc ttacatttgc ttctgggatc cagatctttc
      661 gaagctagcg ctaccggtcg ccaccatggt ctccaagggc gaggagctgt tcaccggggt
      721 ggtgcccatc ctggtcgagc tggacggcga tgtcaacggc cacaagttca gcgtgtccgg
      781 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagctgatct gcaccaccgg
      841 caagctgccc gtgccctggc ccaccctcgt gaccaccctg ggctacggcg tgcagtgctt
      901 cgcccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
      961 ctacgtccag gagcgcacca tcttcttcaa agacgacggc aactacaaga cccgcgccga
     1021 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
     1081 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
     1141 tatcaccgcc gacaagcaga agaacggcat caaggccaac ttcaagatcc gccacaacat
     1201 cgaggacggc ggcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     1261 ccccgtgctg ctgcccgaca accactacct gagctaccag tccaagctga gcaaagaccc
     1321 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     1381 cggcatggac gagctgtaca agggctctgg tgctaccaac ttctcactgt tgaaacaggc
     1441 aggggatgta gaggagaatc cagggcctgg tgctagtgga gactacaaag accatgacgg
     1501 agattataaa gatcatgaca tcgattacaa ggatgacgat gacaagtccg gactcagatc
     1561 tggaggctcc ggaggcccag agccagcgaa gtctgctccc gccccgaaaa agggctccaa
     1621 gaaggcggtg actaaggcgc agaagaaagg cggcaagaag cgcaagcgca gccgcaagga
     1681 gagctattcc atctatgtgt acaaggttct gaagcaggtc caccctgaca ccggcatttc
     1741 gtccaaggcc atgggcatca tgaattcgtt tgtgaacgac attttcgagc gcatcgcagg
     1801 agaggcttcc cgcctggcgc attacaacaa gcgctcgacc atcacctccc gggagatcca
     1861 gacggccgtg cgcctgctgc tgcctgggga gttggccaag cacgccgtgt ccgagggtac
     1921 taaggccatc accaagtaca ccagcgctaa gtaatgaggc gcgccgcggc cgcgaattcg
     1981 atatcataat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     2041 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     2101 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttagttc ttgccacggc
     2161 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga
     2221 caattccgtg gctcgagaga tcttcgactg tgccttctag ttgccagcca tctgttgttt
     2281 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     2341 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     2401 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgag atctcacgtg
     2461 cggaccgagc ggccgcagga acccctagtg atggagttgg ccactccctc tctgcgcgct
     2521 cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg
     2581 gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc
     2641 cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct
     2701 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     2761 ccagcgcctt agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     2821 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     2881 ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct
     2941 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     3001 tccaaactgg aacaacactc aactctatct cgggctattc ttttgattta taagggattt
     3061 tgccgatttc ggtctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     3121 ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg
     3181 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     3241 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     3301 agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat
     3361 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     3421 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     3481 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     3541 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     3601 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     3661 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     3721 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     3781 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     3841 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     3901 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     3961 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     4021 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     4081 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     4141 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     4201 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     4261 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     4321 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     4381 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     4441 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     4501 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     4561 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4621 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4681 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4741 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4801 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4861 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4921 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4981 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     5041 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     5101 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     5161 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgt
//
