LOCUS       sequence-470667-        5547 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0977h 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 5547)
  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 5547)
  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: AiP13461 -
            pAAV-AiE0977h-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5547
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     175..194
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(413..517)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     585..603
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(641..663)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     763..782
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(799..1254)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(936..957)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1148..1167
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   1329..1469
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1340..1469
                     /label=AAV2 ITR
     polyA_signal    1587..1698
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     1885..1905
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1992..2369)
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     CDS             complement(2400..2465)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             complement(2478..2534)
                     /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             complement(join(2544..3254,3255..3257,3258..3260))
                     /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(2586..2607)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2935..2954
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     regulatory      3257..3266
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3376..3385
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     repeat_region   4149..4278
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(4340..4928)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4420..4439)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5099..5547,1..343,344..412))
                     /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"
ORIGIN
        1 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
       61 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
      121 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
      181 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
      241 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
      301 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
      361 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
      421 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
      481 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
      541 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
      601 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
      661 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
      721 cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg
      781 tactgagagt gcaccataaa attgtaaacg ttaatatttt gttaaaattc gcgttaaatt
      841 tttgttaaat cagctcattt tttaaccaat agaccgaaat cggcaaaatc ccttataaat
      901 caaaagaata gcccgagata gagttgagtg ttgttccagt ttggaacaag agtccactat
      961 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     1021 tacgtgaacc atcacccaaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     1081 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     1141 gaaaggaagg gaagaaagcg aaaggagcgg gcgctaaggc gctggcaagt gtagcggtca
     1201 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtactatg
     1261 gttgctttga cgtatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     1321 caggcgcccc tgcaggcagc tgcgcgctcg ctcgctcact gaggccgccc gggcaaagcc
     1381 cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg
     1441 agtggccaac tccatcacta ggggttcctg cggccgctcg gtccgcacgt gagatctcat
     1501 gcctgctatt gtcttcccaa tcctccccct tgctgtcctg ccccacccca ccccccagaa
     1561 tagaatgaca cctactcaga caatgcgatg caatttcctc attttattag gaaaggacag
     1621 tgggagtggc accttccagg gtcaaggaag gcacggggga ggggcaaaca acagatggct
     1681 ggcaactaga aggcacagtc gaagatctct cgagccacgg aattgtcagt gcccaacagc
     1741 cgagcccctg tccagcagcg ggcaaggcag gcggcgatga gttccgccgt ggcaagaact
     1801 aaccaggatt tatacaagga ggagaaaatg aaagccatac gggaagcaat agcatgatac
     1861 aaaggcatta aagcagcgta tccacatagc gtaaaaggag caacatagtt aagaatacca
     1921 gtcaatcttt cacaaatttt gtaatccaga ggttgattat gatatcgaat tcgcggccgc
     1981 ggcgcgcctc attacttagc gctggtgtac ttggtgatgg ccttagtacc ctcggacacg
     2041 gcgtgcttgg ccaactcccc aggcagcagc aggcgcacgg ccgtctggat ctcccgggag
     2101 gtgatggtcg agcgcttgtt gtaatgcgcc aggcgggaag cctctcctgc gatgcgctcg
     2161 aaaatgtcgt tcacaaacga attcatgatg cccatggcct tggacgaaat gccggtgtca
     2221 gggtggacct gcttcagaac cttgtacaca tagatggaat agctctcctt gcggctgcgc
     2281 ttgcgcttct tgccgccttt cttctgcgcc ttagtcaccg ccttcttgga gccctttttc
     2341 ggggcgggag cagacttcgc tggctctggg cctccggagc ctccagatct gagtccggac
     2401 ttgtcatcgt catccttgta atcgatgtca tgatctttat aatctccgtc atggtctttg
     2461 tagtctccac tagcaccagg ccctggattc tcctctacat cccctgcctg tttcaacagt
     2521 gagaagttgg tagcaccaga gcccttgtac agctcgtcca tgccgagagt gatcccggcg
     2581 gcggtcacga actccagcag gaccatgtga tcgcgcttct cgttggggtc tttgctcagc
     2641 ttggactggt agctcaggta gtggttgtcg ggcagcagca cggggccgtc gccgatgggg
     2701 gtgttctgct ggtagtggtc ggcgagctgc acgccgccgt cctcgatgtt gtggcggatc
     2761 ttgaagttgg ccttgatgcc gttcttctgc ttgtcggcgg tgatatagac gttgtggctg
     2821 ttgtagttgt actccagctt gtgccccagg atgttgccgt cctccttgaa gtcgatgccc
     2881 ttcagctcga tgcggttcac cagggtgtcg ccctcgaact tcacctcggc gcgggtcttg
     2941 tagttgccgt cgtctttgaa gaagatggtg cgctcctgga cgtagccttc gggcatggcg
     3001 gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc gggcgaagca ctgcacgccg
     3061 tagcccaggg tggtcacgag ggtgggccag ggcacgggca gcttgccggt ggtgcagatc
     3121 agcttcaggg tcagcttgcc gtaggtggca tcgccctcgc cctcgccgga cacgctgaac
     3181 ttgtggccgt tgacatcgcc gtccagctcg accaggatgg gcaccacccc ggtgaacagc
     3241 tcctcgccct tggagaccat ggtggcgacc ggtagcgcta gcttcgaaag atctggatcc
     3301 cagaagcaaa tgtaagcaat agatggctct gccctgactt ttatgcccag cccgagctcg
     3361 ttgcatctgg ccagaccatg gtggcagggg acgtggaggg aggcctggcg tccctccttc
     3421 cacacgctcc caggcgtctc tgtgcatttg ggggcagcat tcatacttgg ctggtaagaa
     3481 atcctaagga agcttccagc agcattcagc ccccacacct cccatcagtg gagccttcct
     3541 ggggagctgt ctctgcaggc ttgctcaacc tgggctgcag ctcttggcag ctgtacccac
     3601 cgaccctgat gggcccctgc atgtcccact gggtagacag cagctatgaa tactgttcta
     3661 catgtctgct tagatcaagg tatcatctgt tgcaccacca gcagcctgcc cacctcagcc
     3721 agccctccac ctagaacaag agggagcatg gttgctgctg gtgcgtgaca gccctgccaa
     3781 aggttgcaac ttaggagcag agattcaagg cagtgctata aataagccaa gctttatttt
     3841 cctacaggca gcttggtttt tctactgtgt gctgccataa atgtggtaga tatcagatcc
     3901 acctatagac aggcctgcct tgagcctccc atggtgcttc tagccaggta cctttggcaa
     3961 ggtgaccaag ccaggccata gctgggactg tagcatgggg aatgttccag gcactcactc
     4021 aacatcacta gagcagggaa ggcttgctct gagccaaccc ttgttctact gtcttgttcc
     4081 aggtataacc catcagccaa agctccttca atactagctc ttgtctcctg acctacgcgt
     4141 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     4201 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc ggcctcagtg agcgagcgag
     4261 cgcgcagctg cctgcaggac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     4321 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     4381 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     4441 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     4501 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     4561 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     4621 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     4681 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     4741 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     4801 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     4861 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     4921 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     4981 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     5041 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     5101 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     5161 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     5221 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     5281 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     5341 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     5401 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     5461 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     5521 ttagctcctt cggtcctccg atcgttg
//
