LOCUS       sequence-470736-        5080 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0678m 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 5080)
  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 5080)
  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: AiP13659 -
            pAAV-AiE0678m-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5080
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   274..403
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(465..1053)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(545..564)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1224..2015,2016..2084))
                     /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     1847..1866
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2085..2189)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2257..2275
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2313..2335)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2435..2454
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(2471..2926)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2608..2629)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2820..2839
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   3001..3141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3012..3141
                     /label=AAV2 ITR
     polyA_signal    3259..3370
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     3557..3577
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3664..4041)
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     CDS             complement(4072..4137)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             complement(4150..4206)
                     /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(4216..4926,4927..4929,4930..4932))
                     /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(4258..4279)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     4607..4626
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     regulatory      4929..4938
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 gcctatgact gccgaaatgt cttgttctac ttatctctat gaggagcact tagtaagatt
       61 gctttaaata aacatagtgt cttagtcaca gttccatttc tgtgaagagt cattatgacc
      121 aaggcaactc ttatgcaaga aagcatttaa ttggagcctt gtttacagtt tcagaggttt
      181 agttcattgt tatcatggtg gggagcatag cagtacacag gtgggtggtg gagcagtagc
      241 tgaacactac atgatggata cgcgtgcggc cgcaggaacc cctagtgatg gagttggcca
      301 ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc
      361 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggacatgtg agcaaaaggc
      421 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
      481 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
      541 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
      601 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
      661 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
      721 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
      781 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
      841 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
      901 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
      961 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     1021 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     1081 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     1141 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     1201 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     1261 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     1321 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     1381 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     1441 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     1501 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     1561 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     1621 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     1681 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     1741 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     1801 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     1861 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     1921 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     1981 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     2041 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     2101 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     2161 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     2221 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     2281 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     2341 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     2401 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga
     2461 gtgcaccata aaattgtaaa cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     2521 atcagctcat tttttaacca atagaccgaa atcggcaaaa tcccttataa atcaaaagaa
     2581 tagcccgaga tagagttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     2641 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     2701 ccatcaccca aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     2761 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     2821 gggaagaaag cgaaaggagc gggcgctaag gcgctggcaa gtgtagcggt cacgctgcgc
     2881 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtacta tggttgcttt
     2941 gacgtatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
     3001 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
     3061 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
     3121 actccatcac taggggttcc tgcggccgct cggtccgcac gtgagatctc atgcctgcta
     3181 ttgtcttccc aatcctcccc cttgctgtcc tgccccaccc caccccccag aatagaatga
     3241 cacctactca gacaatgcga tgcaatttcc tcattttatt aggaaaggac agtgggagtg
     3301 gcaccttcca gggtcaagga aggcacgggg gaggggcaaa caacagatgg ctggcaacta
     3361 gaaggcacag tcgaagatct ctcgagccac ggaattgtca gtgcccaaca gccgagcccc
     3421 tgtccagcag cgggcaaggc aggcggcgat gagttccgcc gtggcaagaa ctaaccagga
     3481 tttatacaag gaggagaaaa tgaaagccat acgggaagca atagcatgat acaaaggcat
     3541 taaagcagcg tatccacata gcgtaaaagg agcaacatag ttaagaatac cagtcaatct
     3601 ttcacaaatt ttgtaatcca gaggttgatt atgatatcga attcgcggcc gcggcgcgcc
     3661 tcattactta gcgctggtgt acttggtgat ggccttagta ccctcggaca cggcgtgctt
     3721 ggccaactcc ccaggcagca gcaggcgcac ggccgtctgg atctcccggg aggtgatggt
     3781 cgagcgcttg ttgtaatgcg ccaggcggga agcctctcct gcgatgcgct cgaaaatgtc
     3841 gttcacaaac gaattcatga tgcccatggc cttggacgaa atgccggtgt cagggtggac
     3901 ctgcttcaga accttgtaca catagatgga atagctctcc ttgcggctgc gcttgcgctt
     3961 cttgccgcct ttcttctgcg ccttagtcac cgccttcttg gagccctttt tcggggcggg
     4021 agcagacttc gctggctctg ggcctccgga gcctccagat ctgagtccgg acttgtcatc
     4081 gtcatccttg taatcgatgt catgatcttt ataatctccg tcatggtctt tgtagtctcc
     4141 actagcacca ggccctggat tctcctctac atcccctgcc tgtttcaaca gtgagaagtt
     4201 ggtagcacca gagcccttgt acagctcgtc catgccgaga gtgatcccgg cggcggtcac
     4261 gaactccagc aggaccatgt gatcgcgctt ctcgttgggg tctttgctca gcttggactg
     4321 gtagctcagg tagtggttgt cgggcagcag cacggggccg tcgccgatgg gggtgttctg
     4381 ctggtagtgg tcggcgagct gcacgccgcc gtcctcgatg ttgtggcgga tcttgaagtt
     4441 ggccttgatg ccgttcttct gcttgtcggc ggtgatatag acgttgtggc tgttgtagtt
     4501 gtactccagc ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc
     4561 gatgcggttc accagggtgt cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc
     4621 gtcgtctttg aagaagatgg tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa
     4681 gaagtcgtgc tgcttcatgt ggtcggggta gcgggcgaag cactgcacgc cgtagcccag
     4741 ggtggtcacg agggtgggcc agggcacggg cagcttgccg gtggtgcaga tcagcttcag
     4801 ggtcagcttg ccgtaggtgg catcgccctc gccctcgccg gacacgctga acttgtggcc
     4861 gttgacatcg ccgtccagct cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc
     4921 cttggagacc atggtggcga ccggtagcgc tagcttcgaa agatctggat cccagaagca
     4981 aatgtaagca atagatggct ctgccctgac ttttatgccc agcccgagct catacatata
     5041 cactgttcaa aactactcac cagcctatat ttcatcgatg
//
