LOCUS       sequence-470616-        5273 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0946m 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 5273)
  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 5273)
  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: AiP13299 -
            pAAV-AiE0946m-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5273
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     41..60
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(77..532)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(214..235)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     426..445
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   607..747
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   618..747
                     /label=AAV2 ITR
     polyA_signal    865..976
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     1163..1183
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1270..1647)
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     CDS             complement(1678..1743)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             complement(1756..1812)
                     /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(1822..2532,2533..2535,2536..2538))
                     /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(1864..1885)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2213..2232
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     regulatory      2535..2544
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     repeat_region   3153..3282
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(3344..3932)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3424..3443)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4103..4894,4895..4963))
                     /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     4726..4745
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4964..5068)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5136..5154
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5192..5214)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
       61 ctgagagtgc accataaaat tgtaaacgtt aatattttgt taaaattcgc gttaaatttt
      121 tgttaaatca gctcattttt taaccaatag accgaaatcg gcaaaatccc ttataaatca
      181 aaagaatagc ccgagataga gttgagtgtt gttccagttt ggaacaagag tccactatta
      241 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
      301 cgtgaaccat cacccaaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
      361 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
      421 aaggaaggga agaaagcgaa aggagcgggc gctaaggcgc tggcaagtgt agcggtcacg
      481 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtactatggt
      541 tgctttgacg tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
      601 ggcgcccctg caggcagctg cgcgctcgct cgctcactga ggccgcccgg gcaaagcccg
      661 ggcgtcgggc gacctttggt cgcccggcct cagtgagcga gcgagcgcgc agagagggag
      721 tggccaactc catcactagg ggttcctgcg gccgctcggt ccgcacgtga gatctcatgc
      781 ctgctattgt cttcccaatc ctcccccttg ctgtcctgcc ccaccccacc ccccagaata
      841 gaatgacacc tactcagaca atgcgatgca atttcctcat tttattagga aaggacagtg
      901 ggagtggcac cttccagggt caaggaaggc acgggggagg ggcaaacaac agatggctgg
      961 caactagaag gcacagtcga agatctctcg agccacggaa ttgtcagtgc ccaacagccg
     1021 agcccctgtc cagcagcggg caaggcaggc ggcgatgagt tccgccgtgg caagaactaa
     1081 ccaggattta tacaaggagg agaaaatgaa agccatacgg gaagcaatag catgatacaa
     1141 aggcattaaa gcagcgtatc cacatagcgt aaaaggagca acatagttaa gaataccagt
     1201 caatctttca caaattttgt aatccagagg ttgattatga tatcgaattc gcggccgcgg
     1261 cgcgcctcat tacttagcgc tggtgtactt ggtgatggcc ttagtaccct cggacacggc
     1321 gtgcttggcc aactccccag gcagcagcag gcgcacggcc gtctggatct cccgggaggt
     1381 gatggtcgag cgcttgttgt aatgcgccag gcgggaagcc tctcctgcga tgcgctcgaa
     1441 aatgtcgttc acaaacgaat tcatgatgcc catggccttg gacgaaatgc cggtgtcagg
     1501 gtggacctgc ttcagaacct tgtacacata gatggaatag ctctccttgc ggctgcgctt
     1561 gcgcttcttg ccgcctttct tctgcgcctt agtcaccgcc ttcttggagc cctttttcgg
     1621 ggcgggagca gacttcgctg gctctgggcc tccggagcct ccagatctga gtccggactt
     1681 gtcatcgtca tccttgtaat cgatgtcatg atctttataa tctccgtcat ggtctttgta
     1741 gtctccacta gcaccaggcc ctggattctc ctctacatcc cctgcctgtt tcaacagtga
     1801 gaagttggta gcaccagagc ccttgtacag ctcgtccatg ccgagagtga tcccggcggc
     1861 ggtcacgaac tccagcagga ccatgtgatc gcgcttctcg ttggggtctt tgctcagctt
     1921 ggactggtag ctcaggtagt ggttgtcggg cagcagcacg gggccgtcgc cgatgggggt
     1981 gttctgctgg tagtggtcgg cgagctgcac gccgccgtcc tcgatgttgt ggcggatctt
     2041 gaagttggcc ttgatgccgt tcttctgctt gtcggcggtg atatagacgt tgtggctgtt
     2101 gtagttgtac tccagcttgt gccccaggat gttgccgtcc tccttgaagt cgatgccctt
     2161 cagctcgatg cggttcacca gggtgtcgcc ctcgaacttc acctcggcgc gggtcttgta
     2221 gttgccgtcg tctttgaaga agatggtgcg ctcctggacg tagccttcgg gcatggcgga
     2281 cttgaagaag tcgtgctgct tcatgtggtc ggggtagcgg gcgaagcact gcacgccgta
     2341 gcccagggtg gtcacgaggg tgggccaggg cacgggcagc ttgccggtgg tgcagatcag
     2401 cttcagggtc agcttgccgt aggtggcatc gccctcgccc tcgccggaca cgctgaactt
     2461 gtggccgttg acatcgccgt ccagctcgac caggatgggc accaccccgg tgaacagctc
     2521 ctcgcccttg gagaccatgg tggcgaccgg tagcgctagc ttcgaaagat ctggatccca
     2581 gaagcaaatg taagcaatag atggctctgc cctgactttt atgcccagcc cgagctcgtc
     2641 tgtggactta ctgaatgctt tatctactat tatggtgttc catgcaacac tgcttctgac
     2701 caagaactta ctttacaaaa acaaaaacaa aaaaataaaa accatggaga tgggccaaag
     2761 ctcacagcct tcatgggttt accatatgtg ccaccactcg aggcagttag ctgacagaac
     2821 agtgaaatag atttttgaag actcagttac aatgccatat ggctggcaac accttgaagg
     2881 actgtagcaa gcttttccaa aaggctccag atgctagaat cagcacacaa tctgggttgg
     2941 tccttcccta gcatcttctc ccagactcag gaagcaaggg cagacatagg acttccactc
     3001 tgccactagt ttcccgttcc tggggtactt ttctctgctt gtgtaagatt cctcactcct
     3061 ggaaaaaaaa aagggaaaac ttgagaataa ttccttcaaa ctgaaagtta atgtgttcat
     3121 ctgggtgttt gaggctctac gcgtgcggcc gcaggaaccc ctagtgatgg agttggccac
     3181 tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc
     3241 gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggacatgtga gcaaaaggcc
     3301 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     3361 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     3421 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     3481 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     3541 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     3601 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     3661 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     3721 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     3781 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     3841 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     3901 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     3961 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     4021 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     4081 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     4141 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     4201 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     4261 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     4321 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     4381 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     4441 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     4501 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     4561 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     4621 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     4681 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     4741 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     4801 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     4861 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     4921 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     4981 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     5041 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
     5101 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     5161 gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     5221 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcg
//
