LOCUS       sequence-460598-        5142 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0121h_3xC2 is an enhancer sequence, designed to drive 
            AAV-mediated transgene expression in specific brain cell types.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5142)
  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 5142)
  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: AiP12055 -
            pAAV-AiE0121h_3xC2-minCMV-SYFP2-WPR...
FEATURES             Location/Qualifiers
     source          1..5142
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        1054..1092
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      1154..1163
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1160..1162,1163..1165,1166..1876)
                     /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(1205..1226)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1466..1485)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1813..1834
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1969..1989)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2176..2287
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2405..2545
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2405..2534
                     /label=AAV2 ITR
     rep_origin      2620..3075
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2707..2726)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2917..2938
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3092..3111)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3211..3233
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3271..3289)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3357..3461
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3462..3530,3531..4322)
                     /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(3680..3699)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4493..5081
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4982..5001
                     /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 gcgtctatct ggtttataat tagaagtgaa atgattttaa
      181 atggcttcca catggcttag ttttccaagc agttccagtt acatagaact tcatataaac
      241 agataatgag aggctagagt tggagagcac agttcctgct tccaaattgg cttgaagtgg
      301 ggacatctgc tctgcatgaa gcctgggtca ggctgcccct ctcatctgtg acctcctgtg
      361 gtcgggaggg acttgctctg gattatataa aggcttcaca agagtactgg aagcggggac
      421 ggaatcaact tgagcaaaaa acctctatct ggtttataat tagaagtgaa atgattttaa
      481 atggcttcca catggcttag ttttccaagc agttccagtt acatagaact tcatataaac
      541 agataatgag aggctagagt tggagagcac agttcctgct tccaaattgg cttgaagtgg
      601 ggacatctgc tctgcatgaa gcctgggtca ggctgcccct ctcatctgtg acctcctgtg
      661 gtcgggaggg acttgctctg gattatataa aggcttcaca agagtactgg aagcggggac
      721 ggaatcaact tgagcaaaaa acctctatct ggtttataat tagaagtgaa atgattttaa
      781 atggcttcca catggcttag ttttccaagc agttccagtt acatagaact tcatataaac
      841 agataatgag aggctagagt tggagagcac agttcctgct tccaaattgg cttgaagtgg
      901 ggacatctgc tctgcatgaa gcctgggtca ggctgcccct ctcatctgtg acctcctgtg
      961 gtcgggaggg acttgctctg gattatataa aggcttcaca agagtactgg aagcggggac
     1021 ggaatcaact tgagcaaaaa acctgagctc agaggtaggc gtgtacggtg ggaggcctat
     1081 ataagcagag ctggtttagt gaaccgtcag atcgcctggg gatccagatc tttcgaagct
     1141 agcgctaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
     1201 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
     1261 cgagggcgat gccacctacg gcaagctgac cctgaagctg atctgcacca ccggcaagct
     1321 gcccgtgccc tggcccaccc tcgtgaccac cctgggctac ggcgtgcagt gcttcgcccg
     1381 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
     1441 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
     1501 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     1561 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcac
     1621 cgccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga
     1681 cggcggcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     1741 gctgctgccc gacaaccact acctgagcta ccagtccaag ctgagcaaag accccaacga
     1801 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     1861 ggacgagctg tacaagtaag tcgacggcgc gccgcggccg cgaattcgat atcataatca
     1921 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     1981 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     2041 tttcattttc tcctccttgt ataaatcctg gttagttctt gccacggcgg aactcatcgc
     2101 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggc
     2161 tcgagagatc ttcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     2221 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     2281 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     2341 gcaaggggga ggattgggaa gacaatagca ggcatgagat ctcacgtgcg gaccgagcgg
     2401 ccgcaggaac ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact
     2461 gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc
     2521 gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg
     2581 tgcggtattt cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat
     2641 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccttag
     2701 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     2761 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     2821 ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     2881 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
     2941 caacactcaa ctctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg
     3001 tctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat
     3061 taacgtttac aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     3121 gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     3181 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     3241 cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta
     3301 atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg
     3361 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     3421 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     3481 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     3541 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     3601 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     3661 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     3721 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     3781 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     3841 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     3901 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     3961 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     4021 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     4081 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     4141 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     4201 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     4261 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     4321 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     4381 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     4441 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     4501 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     4561 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     4621 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     4681 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     4741 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     4801 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     4861 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     4921 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     4981 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     5041 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     5101 ttttacggtt cctggccttt tgctggcctt ttgctcacat gt
//
