LOCUS       sequence-470634-        5387 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0969m 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 5387)
  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 5387)
  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: AiP13315 -
            pAAV-AiE0969m-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5387
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    157..268
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     455..475
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(562..939)
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     CDS             complement(970..1035)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             complement(1048..1104)
                     /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(1114..1824,1825..1827,1828..1830))
                     /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(1156..1177)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1505..1524
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     regulatory      1827..1836
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     repeat_region   2559..2688
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(2750..3338)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2830..2849)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3509..4300,4301..4369))
                     /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     4132..4151
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4370..4474)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4542..4560
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4598..4620)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4720..4739
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(4756..5211)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4893..4914)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5105..5124
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   join(5286..5387,1..39)
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   join(5297..5387,1..39)
                     /label=AAV2 ITR
ORIGIN
        1 gcagagaggg agtggccaac tccatcacta ggggttcctg cggccgctcg gtccgcacgt
       61 gagatctcat gcctgctatt gtcttcccaa tcctccccct tgctgtcctg ccccacccca
      121 ccccccagaa tagaatgaca cctactcaga caatgcgatg caatttcctc attttattag
      181 gaaaggacag tgggagtggc accttccagg gtcaaggaag gcacggggga ggggcaaaca
      241 acagatggct ggcaactaga aggcacagtc gaagatctct cgagccacgg aattgtcagt
      301 gcccaacagc cgagcccctg tccagcagcg ggcaaggcag gcggcgatga gttccgccgt
      361 ggcaagaact aaccaggatt tatacaagga ggagaaaatg aaagccatac gggaagcaat
      421 agcatgatac aaaggcatta aagcagcgta tccacatagc gtaaaaggag caacatagtt
      481 aagaatacca gtcaatcttt cacaaatttt gtaatccaga ggttgattat gatatcgaat
      541 tcgcggccgc ggcgcgcctc attacttagc gctggtgtac ttggtgatgg ccttagtacc
      601 ctcggacacg gcgtgcttgg ccaactcccc aggcagcagc aggcgcacgg ccgtctggat
      661 ctcccgggag gtgatggtcg agcgcttgtt gtaatgcgcc aggcgggaag cctctcctgc
      721 gatgcgctcg aaaatgtcgt tcacaaacga attcatgatg cccatggcct tggacgaaat
      781 gccggtgtca gggtggacct gcttcagaac cttgtacaca tagatggaat agctctcctt
      841 gcggctgcgc ttgcgcttct tgccgccttt cttctgcgcc ttagtcaccg ccttcttgga
      901 gccctttttc ggggcgggag cagacttcgc tggctctggg cctccggagc ctccagatct
      961 gagtccggac ttgtcatcgt catccttgta atcgatgtca tgatctttat aatctccgtc
     1021 atggtctttg tagtctccac tagcaccagg ccctggattc tcctctacat cccctgcctg
     1081 tttcaacagt gagaagttgg tagcaccaga gcccttgtac agctcgtcca tgccgagagt
     1141 gatcccggcg gcggtcacga actccagcag gaccatgtga tcgcgcttct cgttggggtc
     1201 tttgctcagc ttggactggt agctcaggta gtggttgtcg ggcagcagca cggggccgtc
     1261 gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc acgccgccgt cctcgatgtt
     1321 gtggcggatc ttgaagttgg ccttgatgcc gttcttctgc ttgtcggcgg tgatatagac
     1381 gttgtggctg ttgtagttgt actccagctt gtgccccagg atgttgccgt cctccttgaa
     1441 gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg ccctcgaact tcacctcggc
     1501 gcgggtcttg tagttgccgt cgtctttgaa gaagatggtg cgctcctgga cgtagccttc
     1561 gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc gggcgaagca
     1621 ctgcacgccg tagcccaggg tggtcacgag ggtgggccag ggcacgggca gcttgccggt
     1681 ggtgcagatc agcttcaggg tcagcttgcc gtaggtggca tcgccctcgc cctcgccgga
     1741 cacgctgaac ttgtggccgt tgacatcgcc gtccagctcg accaggatgg gcaccacccc
     1801 ggtgaacagc tcctcgccct tggagaccat ggtggcgacc ggtagcgcta gcttcgaaag
     1861 atctggatcc cagaagcaaa tgtaagcaat agatggctct gccctgactt ttatgcccag
     1921 cccgagctct ccatttcctt ccttcccagc agccacttcc tgcgtgattt acacagctga
     1981 acgtagcctt tttcttatcc ccatttcaaa cctggaggca tctgaacaca tctcacagtg
     2041 ttgatggaag catcactatc ttcttcctta cctggagaaa ttgctgtgga gggctgtgtg
     2101 aaatattctc ttgataatta gggtttattt ttaaagcttg ttttttaaag ttttcataca
     2161 agaatataat gtgtgtcctg cctactcttc ctcataatcc ctcccccctt tcccttttct
     2221 ctcctatgag tccccgtggc cacctacata ctttcccttc tatatttatg ccatctgtga
     2281 atatataact ttgtgtagct atataaaaat ctacgaatct ttctgagatt ggcacaattg
     2341 gcttaaaatt tatttctggt tgcattattt tccagaaaat tactcactct cgttcttctt
     2401 gggtatgttt cattggatgt accacatcta gctatatgta tctatgtgta tatgtttatc
     2461 catctatatc tgtagacata tatgctataa tagggtattc actttagacc actttccttc
     2521 atcattcctg aaggggagtt accaacgcgt gcggccgcag gaacccctag tgatggagtt
     2581 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     2641 acgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcaggac atgtgagcaa
     2701 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     2761 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     2821 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     2881 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     2941 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     3001 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     3061 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     3121 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     3181 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     3241 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     3301 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     3361 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     3421 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     3481 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     3541 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     3601 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     3661 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     3721 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     3781 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     3841 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     3901 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     3961 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     4021 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     4081 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     4141 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     4201 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     4261 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     4321 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     4381 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     4441 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     4501 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     4561 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     4621 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     4681 cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac
     4741 tgagagtgca ccataaaatt gtaaacgtta atattttgtt aaaattcgcg ttaaattttt
     4801 gttaaatcag ctcatttttt aaccaataga ccgaaatcgg caaaatccct tataaatcaa
     4861 aagaatagcc cgagatagag ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     4921 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     4981 gtgaaccatc acccaaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     5041 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     5101 aggaagggaa gaaagcgaaa ggagcgggcg ctaaggcgct ggcaagtgta gcggtcacgc
     5161 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tactatggtt
     5221 gctttgacgt atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
     5281 gcgcccctgc aggcagctgc gcgctcgctc gctcactgag gccgcccggg caaagcccgg
     5341 gcgtcgggcg acctttggtc gcccggcctc agtgagcgag cgagcgc
//
