LOCUS       sequence-461875-        5203 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0963m 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 5203)
  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 5203)
  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: AiP13301 -
            pAAV-AiE0963m-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5203
                     /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      669..678
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(675..677,678..680,681..1391)
                     /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(981..1000)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1328..1349
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1401..1457
                     /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             1470..1535
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1566..1943
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(2030..2050)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2237..2348
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2466..2606
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2466..2595
                     /label=AAV2 ITR
     rep_origin      2681..3136
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2768..2787)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2978..2999
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3153..3172)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3272..3294
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3332..3350)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3418..3522
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3523..3591,3592..4383)
                     /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(3741..3760)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4554..5142
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5043..5062
                     /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 gcgtatcttc agggtcacag gcacataaag gtggaagggg
      181 agaacatatg cagtccatct gtctgtctgt ctttgcacat aggcctcttt accaagccac
      241 cctcggctcg atctccctgc acgcaaccta ccccacttag tagatcttca accctgatgg
      301 catcatcacg gtttctgtcc cctgacctct gctccgagct taccccactt gtctccattg
      361 cagtgcctac agcgctcagc ccgtatgtgc cttgcacagc ctctgtgtgc cgtgcgcagc
      421 atcagtgtgc cgtgcgcagc ctctgtccac gtttacagtg ggtgttacca gcattcctcc
      481 tgctttatca tcctctgtgt tgatgcggag gtcaacacat ccttggctgc tatgagaaca
      541 gaaggacaca gtcagtggac tttgtgggac ttggagagct cgggctgggc ataaaagtca
      601 gggcagagcc atctattgct tacatttgct tctgggatcc agatctttcg aagctagcgc
      661 taccggtcgc caccatggtc tccaagggcg aggagctgtt caccggggtg gtgcccatcc
      721 tggtcgagct ggacggcgat gtcaacggcc acaagttcag cgtgtccggc gagggcgagg
      781 gcgatgccac ctacggcaag ctgaccctga agctgatctg caccaccggc aagctgcccg
      841 tgccctggcc caccctcgtg accaccctgg gctacggcgt gcagtgcttc gcccgctacc
      901 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
      961 agcgcaccat cttcttcaaa gacgacggca actacaagac ccgcgccgag gtgaagttcg
     1021 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     1081 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcaccgccg
     1141 acaagcagaa gaacggcatc aaggccaact tcaagatccg ccacaacatc gaggacggcg
     1201 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     1261 tgcccgacaa ccactacctg agctaccagt ccaagctgag caaagacccc aacgagaagc
     1321 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     1381 agctgtacaa gggctctggt gctaccaact tctcactgtt gaaacaggca ggggatgtag
     1441 aggagaatcc agggcctggt gctagtggag actacaaaga ccatgacgga gattataaag
     1501 atcatgacat cgattacaag gatgacgatg acaagtccgg actcagatct ggaggctccg
     1561 gaggcccaga gccagcgaag tctgctcccg ccccgaaaaa gggctccaag aaggcggtga
     1621 ctaaggcgca gaagaaaggc ggcaagaagc gcaagcgcag ccgcaaggag agctattcca
     1681 tctatgtgta caaggttctg aagcaggtcc accctgacac cggcatttcg tccaaggcca
     1741 tgggcatcat gaattcgttt gtgaacgaca ttttcgagcg catcgcagga gaggcttccc
     1801 gcctggcgca ttacaacaag cgctcgacca tcacctcccg ggagatccag acggccgtgc
     1861 gcctgctgct gcctggggag ttggccaagc acgccgtgtc cgagggtact aaggccatca
     1921 ccaagtacac cagcgctaag taatgaggcg cgccgcggcc gcgaattcga tatcataatc
     1981 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     2041 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     2101 ctttcatttt ctcctccttg tataaatcct ggttagttct tgccacggcg gaactcatcg
     2161 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     2221 ctcgagagat cttcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     2281 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     2341 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     2401 agcaaggggg aggattggga agacaatagc aggcatgaga tctcacgtgc ggaccgagcg
     2461 gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac
     2521 tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag
     2581 cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct
     2641 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
     2701 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta
     2761 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     2821 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     2881 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     2941 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3001 acaacactca actctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     3061 gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     3121 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3181 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     3241 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     3301 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     3361 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     3421 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     3481 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     3541 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     3601 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     3661 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     3721 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     3781 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     3841 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     3901 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     3961 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     4021 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     4081 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     4141 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     4201 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     4261 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     4321 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     4381 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     4441 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4501 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4561 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     4621 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     4681 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     4741 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     4801 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     4861 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     4921 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     4981 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     5041 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     5101 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     5161 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgt
//
