LOCUS       sequence-470658-        5116 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0972h 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 5116)
  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 5116)
  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: AiP13456 -
            pAAV-AiE0972h-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5116
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      605..1193
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1094..1113
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1255..1384
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      1836..1845
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1842..1844,1845..1847,1848..2558)
                     /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(2148..2167)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2495..2516
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2568..2624
                     /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             2637..2702
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             2733..3110
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(3197..3217)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    3404..3515
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3633..3773
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3633..3762
                     /label=AAV2 ITR
     rep_origin      3848..4303
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3935..3954)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4145..4166
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4320..4339)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4439..4461
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4499..4517)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4585..4689
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4690..4758,4759..5116,1..434)
                     /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(4908..4927)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
       61 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
      121 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
      181 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
      241 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
      301 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
      361 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
      421 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
      481 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
      541 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
      601 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
      661 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
      721 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
      781 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
      841 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
      901 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
      961 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     1021 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     1081 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     1141 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     1201 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgtcctgca
     1261 ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc gtcgggcgac ctttggtcgc
     1321 ccggcctcag tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt
     1381 tcctgcggcc gcacgcgtca ggtcagctgg tccttgccca ccgagctcct ctggacccac
     1441 ccctcttttt gggcccgcac ccatctggct cctcccaccc atctggcccc acccactccc
     1501 ctgctttcca tccctgcctt gccctgtgac tggccccacc ctccatccct tttggtccct
     1561 ccttgcctgc cgagctcctc tggacccacc cctctttcag ttggggcccc gccctcattt
     1621 ggcccctttc acccgtctgg ccctgcccat tcccaccttc caccctggcc ccgccctgtg
     1681 gcagacccca ttctccattc cttgtggccc ttctcctctg gaccaacccc tctgtccatt
     1741 aggagctcgg gctgggcata aaagtcaggg cagagccatc tattgcttac atttgcttct
     1801 gggatccaga tctttcgaag ctagcgctac cggtcgccac catggtctcc aagggcgagg
     1861 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgatgtc aacggccaca
     1921 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagc
     1981 tgatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgggct
     2041 acggcgtgca gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     2101 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaagac gacggcaact
     2161 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     2221 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     2281 acagccacaa cgtctatatc accgccgaca agcagaagaa cggcatcaag gccaacttca
     2341 agatccgcca caacatcgag gacggcggcg tgcagctcgc cgaccactac cagcagaaca
     2401 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc taccagtcca
     2461 agctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     2521 ccgccgggat cactctcggc atggacgagc tgtacaaggg ctctggtgct accaacttct
     2581 cactgttgaa acaggcaggg gatgtagagg agaatccagg gcctggtgct agtggagact
     2641 acaaagacca tgacggagat tataaagatc atgacatcga ttacaaggat gacgatgaca
     2701 agtccggact cagatctgga ggctccggag gcccagagcc agcgaagtct gctcccgccc
     2761 cgaaaaaggg ctccaagaag gcggtgacta aggcgcagaa gaaaggcggc aagaagcgca
     2821 agcgcagccg caaggagagc tattccatct atgtgtacaa ggttctgaag caggtccacc
     2881 ctgacaccgg catttcgtcc aaggccatgg gcatcatgaa ttcgtttgtg aacgacattt
     2941 tcgagcgcat cgcaggagag gcttcccgcc tggcgcatta caacaagcgc tcgaccatca
     3001 cctcccggga gatccagacg gccgtgcgcc tgctgctgcc tggggagttg gccaagcacg
     3061 ccgtgtccga gggtactaag gccatcacca agtacaccag cgctaagtaa tgaggcgcgc
     3121 cgcggccgcg aattcgatat cataatcaac ctctggatta caaaatttgt gaaagattga
     3181 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt
     3241 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt
     3301 tagttcttgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     3361 ggctgttggg cactgacaat tccgtggctc gagagatctt cgactgtgcc ttctagttgc
     3421 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     3481 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     3541 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     3601 catgagatct cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac
     3661 tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc
     3721 gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct
     3781 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa
     3841 ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     3901 gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt
     3961 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     4021 cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt
     4081 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     4141 aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt
     4201 gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa
     4261 aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt
     4321 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     4381 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     4441 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     4501 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     4561 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     4621 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     4681 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     4741 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     4801 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     4861 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     4921 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     4981 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5041 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     5101 acaacgatcg gaggac
//
