LOCUS       sequence-462824-        5592 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0140h_3xC2 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 5592)
  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 5592)
  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: AiP13400 -
            pAAV-AiE0140h_3xC2-minBG-SYFP2-P2A-...
FEATURES             Location/Qualifiers
     source          1..5592
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      838..847
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(844..846,847..849,850..1560)
                     /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(1150..1169)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1497..1518
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1570..1626
                     /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             1639..1704
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1735..2112
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(2419..2439)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2626..2737
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2855..2995
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2855..2984
                     /label=AAV2 ITR
     rep_origin      3070..3525
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3157..3176)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3367..3388
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3542..3561)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3661..3683
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3721..3739)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3807..3911
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3912..3980,3981..4772)
                     /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(4130..4149)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4943..5531
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5432..5451
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtccaaga aattagaata aagggaaaaa tgactttgag
      181 atgtactttc ttaaaataac tgaagtaatg accgcatgag ccagctgatg ggtttttaaa
      241 tggatacgtt tctatcagcc tgctgcttga tgccagagcc aaatatatat ggagtgtatt
      301 atatcatatc ctgtaggcag gagactgttg gggcattggg ggacccaaga aattagaata
      361 aagggaaaaa tgactttgag atgtactttc ttaaaataac tgaagtaatg accgcatgag
      421 ccagctgatg ggtttttaaa tggatacgtt tctatcagcc tgctgcttga tgccagagcc
      481 aaatatatat ggagtgtatt atatcatatc ctgtaggcag gagactgttg gggcattggg
      541 ggacccaaga aattagaata aagggaaaaa tgactttgag atgtactttc ttaaaataac
      601 tgaagtaatg accgcatgag ccagctgatg ggtttttaaa tggatacgtt tctatcagcc
      661 tgctgcttga tgccagagcc aaatatatat ggagtgtatt atatcatatc ctgtaggcag
      721 gagactgttg gggcattggg ggacgagctc gggctgggca taaaagtcag ggcagagcca
      781 tctattgctt acatttgctt ctgggatcca gatctttcga agctagcgct accggtcgcc
      841 accatggtct ccaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      901 gacggcgatg tcaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      961 tacggcaagc tgaccctgaa gctgatctgc accaccggca agctgcccgt gccctggccc
     1021 accctcgtga ccaccctggg ctacggcgtg cagtgcttcg cccgctaccc cgaccacatg
     1081 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     1141 ttcttcaaag acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     1201 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1261 cacaagctgg agtacaacta caacagccac aacgtctata tcaccgccga caagcagaag
     1321 aacggcatca aggccaactt caagatccgc cacaacatcg aggacggcgg cgtgcagctc
     1381 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     1441 cactacctga gctaccagtc caagctgagc aaagacccca acgagaagcg cgatcacatg
     1501 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     1561 ggctctggtg ctaccaactt ctcactgttg aaacaggcag gggatgtaga ggagaatcca
     1621 gggcctggtg ctagtggaga ctacaaagac catgacggag attataaaga tcatgacatc
     1681 gattacaagg atgacgatga caagtccgga ctcagatctg gaggctccgg aggcccagag
     1741 ccagcgaagt ctgctcccgc cccgaaaaag ggctccaaga aggcggtgac taaggcgcag
     1801 aagaaaggcg gcaagaagcg caagcgcagc cgcaaggaga gctattccat ctatgtgtac
     1861 aaggttctga agcaggtcca ccctgacacc ggcatttcgt ccaaggccat gggcatcatg
     1921 aattcgtttg tgaacgacat tttcgagcgc atcgcaggag aggcttcccg cctggcgcat
     1981 tacaacaagc gctcgaccat cacctcccgg gagatccaga cggccgtgcg cctgctgctg
     2041 cctggggagt tggccaagca cgccgtgtcc gagggtacta aggccatcac caagtacacc
     2101 agcgctaagt aatgaggcgc gccgcggcct taaagagacc ggttcactgt gacagtaaaa
     2161 gagaccggtt cactgtgaga atgaaagaga ccggttcact gtgatcggaa aagagaccgg
     2221 ttcactgtga gcggccttga aacccagcag acaatgtagc tcagtagaaa cccagcagac
     2281 aatgtagctg aatggaaacc cagcagacaa tgtagcttcg gagaaaccca gcagacaatg
     2341 tagctgtcga cgaattcgat atcataatca acctctggat tacaaaattt gtgaaagatt
     2401 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     2461 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     2521 gttagttctt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     2581 tcggctgttg ggcactgaca attccgtggc tcgagagatc ttcgactgtg ccttctagtt
     2641 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     2701 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     2761 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     2821 ggcatgagat ctcacgtgcg gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc
     2881 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     2941 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc
     3001 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc
     3061 aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     3121 gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc tttcgctttc ttcccttcct
     3181 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     3241 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac
     3301 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     3361 ttaatagtgg actcttgttc caaactggaa caacactcaa ctctatctcg ggctattctt
     3421 ttgatttata agggattttg ccgatttcgg tctattggtt aaaaaatgag ctgatttaac
     3481 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca
     3541 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     3601 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     3661 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     3721 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     3781 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     3841 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     3901 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     3961 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     4021 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     4081 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     4141 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     4201 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     4261 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     4321 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     4381 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     4441 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     4501 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     4561 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     4621 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     4681 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     4741 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     4801 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     4861 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     4921 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     4981 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     5041 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     5101 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     5161 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     5221 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     5281 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     5341 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     5401 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     5461 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     5521 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     5581 ttgctcacat gt
//
