LOCUS       sequence-474125-        5363 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE1014h_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 5363)
  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 5363)
  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: AiP14282 -
            pAAV-AiE1014h_3xC2-minBG-SYFP2-P2A-...
FEATURES             Location/Qualifiers
     source          1..5363
                     /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      829..838
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(835..837,838..840,841..1551)
                     /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(1141..1160)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1488..1509
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1561..1617
                     /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             1630..1695
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1726..2103
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(2190..2210)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2397..2508
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2626..2766
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2626..2755
                     /label=AAV2 ITR
     rep_origin      2841..3296
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2928..2947)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3138..3159
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3313..3332)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3432..3454
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3492..3510)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3578..3682
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3683..3751,3752..4543)
                     /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(3901..3920)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4714..5302
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5203..5222
                     /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 gcgtaattca gaaagagtat acaaaacatc tgaagagtgc
      181 tctatttagc gttatcatag gcccttagaa taacaaactt cagagagagc tatatagtgc
      241 agctgagaac tgtctgttct tcgaacagga gtcatcactc tacacttagc tgaattttta
      301 atattcctgt aatcatctgc aaaatatact tgtgcacatt taattcagaa agagtataca
      361 aaacatctga agagtgctct atttagcgtt atcataggcc cttagaataa caaacttcag
      421 agagagctat atagtgcagc tgagaactgt ctgttcttcg aacaggagtc atcactctac
      481 acttagctga atttttaata ttcctgtaat catctgcaaa atatacttgt gcacatttaa
      541 ttcagaaaga gtatacaaaa catctgaaga gtgctctatt tagcgttatc ataggccctt
      601 agaataacaa acttcagaga gagctatata gtgcagctga gaactgtctg ttcttcgaac
      661 aggagtcatc actctacact tagctgaatt tttaatattc ctgtaatcat ctgcaaaata
      721 tacttgtgca catttgagct cgggctgggc ataaaagtca gggcagagcc atctattgct
      781 tacatttgct tctgggatcc agatctttcg aagctagcgc taccggtcgc caccatggtc
      841 tccaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgat
      901 gtcaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
      961 ctgaccctga agctgatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     1021 accaccctgg gctacggcgt gcagtgcttc gcccgctacc ccgaccacat gaagcagcac
     1081 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaaa
     1141 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     1201 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     1261 gagtacaact acaacagcca caacgtctat atcaccgccg acaagcagaa gaacggcatc
     1321 aaggccaact tcaagatccg ccacaacatc gaggacggcg gcgtgcagct cgccgaccac
     1381 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     1441 agctaccagt ccaagctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     1501 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gggctctggt
     1561 gctaccaact tctcactgtt gaaacaggca ggggatgtag aggagaatcc agggcctggt
     1621 gctagtggag actacaaaga ccatgacgga gattataaag atcatgacat cgattacaag
     1681 gatgacgatg acaagtccgg actcagatct ggaggctccg gaggcccaga gccagcgaag
     1741 tctgctcccg ccccgaaaaa gggctccaag aaggcggtga ctaaggcgca gaagaaaggc
     1801 ggcaagaagc gcaagcgcag ccgcaaggag agctattcca tctatgtgta taaggttctg
     1861 aagcaggtcc accctgacac cggcatttcg tccaaggcca tgggcatcat gaactcgttt
     1921 gtgaacgaca ttttcgagcg catcgcaggt gaggcttccc gcctggcgca ttacaacaag
     1981 cgctcgacca tcacctccag ggagatccag acggccgtgc gcctgctgct gcctggggag
     2041 ttggccaagc acgccgtgtc cgagggtact aaggccatca ccaagtacac cagcgctaag
     2101 taatgaggcg cgccgcggcc gcgaattcga tatcataatc aacctctgga ttacaaaatt
     2161 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     2221 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     2281 tataaatcct ggttagttct tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
     2341 tggacagggg ctcggctgtt gggcactgac aattccgtgg ctcgagagat cttcgactgt
     2401 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     2461 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     2521 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     2581 agacaatagc aggcatgaga tctcacgtgc ggaccgagcg gccgcaggaa cccctagtga
     2641 tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg
     2701 tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc
     2761 tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     2821 tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     2881 ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt
     2941 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     3001 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg
     3061 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     3121 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca actctatctc
     3181 gggctattct tttgatttat aagggatttt gccgatttcg gtctattggt taaaaaatga
     3241 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg
     3301 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc
     3361 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     3421 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     3481 gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
     3541 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     3601 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     3661 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     3721 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     3781 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     3841 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     3901 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     3961 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     4021 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     4081 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     4141 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     4201 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     4261 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     4321 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     4381 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     4441 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     4501 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     4561 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     4621 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     4681 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     4741 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     4801 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     4861 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     4921 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     4981 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     5041 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     5101 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     5161 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     5221 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     5281 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     5341 ttgctggcct tttgctcaca tgt
//
