LOCUS       sequence-462739-        5067 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0050m_3xC2 is an enhancer sequence, designed to drive 
            AAV-mediated transgene expression in cortical L4/5_IT neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5067)
  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 5067)
  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: AiP12211 -
            pAAV-AiE0050m_3xC2-minCMV-SYFP2-WPR...
FEATURES             Location/Qualifiers
     source          1..5067
                     /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        979..1017
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      1079..1088
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1085..1087,1088..1090,1091..1801)
                     /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(1130..1151)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1391..1410)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1738..1759
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1894..1914)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2101..2212
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2330..2470
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2330..2459
                     /label=AAV2 ITR
     rep_origin      2545..3000
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2632..2651)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2842..2863
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3017..3036)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3136..3158
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3196..3214)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3282..3386
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3387..3455,3456..4247)
                     /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(3605..3624)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4418..5006
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4907..4926
                     /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 gcgtaggcaa cttgttaaaa atctcagtca aacttacata
      181 ctatatagaa cagcatggtg aatttagggc acatggatat aaaatggaag tttcttattc
      241 agtagcagca acttgtgggc acaggagttg gcaaagataa aaatgtccaa agtcacaaat
      301 acaatgtata gttagtcata ggtgctgtta tttgcctcaa aaaatagact tttattttgc
      361 ctttcttttc tttaaccaca ctcaaaatta gagaacagag acaaaaccca gcaggaaata
      421 ggcaacttgt taaaaatctc agtcaaactt acatactata tagaacagca tggtgaattt
      481 agggcacatg gatataaaat ggaagtttct tattcagtag cagcaacttg tgggcacagg
      541 agttggcaaa gataaaaatg tccaaagtca caaatacaat gtatagttag tcataggtgc
      601 tgttatttgc ctcaaaaaat agacttttat tttgcctttc ttttctttaa ccacactcaa
      661 aattagagaa cagagacaaa acccagcagg aaataggcaa cttgttaaaa atctcagtca
      721 aacttacata ctatatagaa cagcatggtg aatttagggc acatggatat aaaatggaag
      781 tttcttattc agtagcagca acttgtgggc acaggagttg gcaaagataa aaatgtccaa
      841 agtcacaaat acaatgtata gttagtcata ggtgctgtta tttgcctcaa aaaatagact
      901 tttattttgc ctttcttttc tttaaccaca ctcaaaatta gagaacagag acaaaaccca
      961 gcaggaaatg agctcagagg taggcgtgta cggtgggagg cctatataag cagagctggt
     1021 ttagtgaacc gtcagatcgc ctggggatcc agatctttcg aagctagcgc taccggtcgc
     1081 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     1141 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     1201 ctacggcaag ctgaccctga agctgatctg caccaccggc aagctgcccg tgccctggcc
     1261 caccctcgtg accaccctgg gctacggcgt gcagtgcttc gcccgctacc ccgaccacat
     1321 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     1381 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1441 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1501 gcacaagctg gagtacaact acaacagcca caacgtctat atcaccgccg acaagcagaa
     1561 gaacggcatc aaggccaact tcaagatccg ccacaacatc gaggacggcg gcgtgcagct
     1621 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1681 ccactacctg agctaccagt ccaagctgag caaagacccc aacgagaagc gcgatcacat
     1741 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     1801 gtaagtcgac ggcgcgccgc ggccgcgaat tcgatatcat aatcaacctc tggattacaa
     1861 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata
     1921 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc
     1981 cttgtataaa tcctggttag ttcttgccac ggcggaactc atcgccgcct gccttgcccg
     2041 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggctcgag agatcttcga
     2101 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     2161 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     2221 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     2281 gggaagacaa tagcaggcat gagatctcac gtgcggaccg agcggccgca ggaaccccta
     2341 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca
     2401 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc
     2461 tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     2521 cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     2581 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cttagcgccc gctcctttcg
     2641 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     2701 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     2761 tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     2821 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta
     2881 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa
     2941 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     3001 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     3061 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     3121 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     3181 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     3241 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     3301 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     3361 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     3421 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3481 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3541 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3601 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     3661 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3721 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3781 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3841 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3901 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     3961 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     4021 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     4081 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     4141 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     4201 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     4261 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     4321 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     4381 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4441 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4501 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4561 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4621 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4681 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4741 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4801 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4861 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4921 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4981 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     5041 ccttttgctg gccttttgct cacatgt
//
