LOCUS       sequence-470637-        5184 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0875m 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 5184)
  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 5184)
  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: AiP13367 -
            pAAV-AiE0875m-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5184
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(57..76)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      870..1458
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1359..1378
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1520..1649
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      2169..2178
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2175..2177,2178..2180,2181..2891)
                     /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(2481..2500)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2828..2849
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2901..2957
                     /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             2970..3035
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3066..3443
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(3530..3550)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    3737..3848
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3966..4106
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3966..4095
                     /label=AAV2 ITR
     rep_origin      4181..4636
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4268..4287)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4478..4499
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4653..4672)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4772..4794
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4832..4850)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4918..5022
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5023..5091,5092..5184,1..699)
                     /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"
ORIGIN
        1 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
       61 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
      121 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
      181 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
      241 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
      301 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
      361 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
      421 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
      481 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
      541 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
      601 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
      661 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
      721 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
      781 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
      841 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
      901 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
      961 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     1021 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     1081 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     1141 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     1201 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     1261 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     1321 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     1381 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     1441 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     1501 tggccttttg ctcacatgtc ctgcaggcag ctgcgcgctc gctcgctcac tgaggccgcc
     1561 cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg
     1621 agtggccaac tccatcacta ggggttcctg cggccgcacg cgtactgact gaggctgctc
     1681 aatgtaccat attaaaaagg gagctgcggc taagccagtt tctgcaggac agagccacag
     1741 actatgggat acttccaagg agaaggaagg agtggcgagg tggcactgcg tcagctgacc
     1801 gctagactgt gactttatcc agctctgtgt tagctgagtg atgacagtaa tgccgtatct
     1861 acacacccac cgcctactgg gaacctccac atgagtacat aacatagcat ctgaggccag
     1921 cagtgcgcag aacacatctg ctcatctaca gtgccttcaa acgggaggga agtaccagtc
     1981 ctctctcagt gagaacacca tccgtcctgg gtgagagctt taggcattgc catggctccc
     2041 tgtcactcgg ccctgaccca cacattcaca gccatgagct cgggctgggc ataaaagtca
     2101 gggcagagcc atctattgct tacatttgct tctgggatcc agatctttcg aagctagcgc
     2161 taccggtcgc caccatggtc tccaagggcg aggagctgtt caccggggtg gtgcccatcc
     2221 tggtcgagct ggacggcgat gtcaacggcc acaagttcag cgtgtccggc gagggcgagg
     2281 gcgatgccac ctacggcaag ctgaccctga agctgatctg caccaccggc aagctgcccg
     2341 tgccctggcc caccctcgtg accaccctgg gctacggcgt gcagtgcttc gcccgctacc
     2401 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
     2461 agcgcaccat cttcttcaaa gacgacggca actacaagac ccgcgccgag gtgaagttcg
     2521 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     2581 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcaccgccg
     2641 acaagcagaa gaacggcatc aaggccaact tcaagatccg ccacaacatc gaggacggcg
     2701 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     2761 tgcccgacaa ccactacctg agctaccagt ccaagctgag caaagacccc aacgagaagc
     2821 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     2881 agctgtacaa gggctctggt gctaccaact tctcactgtt gaaacaggca ggggatgtag
     2941 aggagaatcc agggcctggt gctagtggag actacaaaga ccatgacgga gattataaag
     3001 atcatgacat cgattacaag gatgacgatg acaagtccgg actcagatct ggaggctccg
     3061 gaggcccaga gccagcgaag tctgctcccg ccccgaaaaa gggctccaag aaggcggtga
     3121 ctaaggcgca gaagaaaggc ggcaagaagc gcaagcgcag ccgcaaggag agctattcca
     3181 tctatgtgta caaggttctg aagcaggtcc accctgacac cggcatttcg tccaaggcca
     3241 tgggcatcat gaattcgttt gtgaacgaca ttttcgagcg catcgcagga gaggcttccc
     3301 gcctggcgca ttacaacaag cgctcgacca tcacctcccg ggagatccag acggccgtgc
     3361 gcctgctgct gcctggggag ttggccaagc acgccgtgtc cgagggtact aaggccatca
     3421 ccaagtacac cagcgctaag taatgaggcg cgccgcggcc gcgaattcga tatcataatc
     3481 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     3541 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     3601 ctttcatttt ctcctccttg tataaatcct ggttagttct tgccacggcg gaactcatcg
     3661 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     3721 ctcgagagat cttcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     3781 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     3841 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     3901 agcaaggggg aggattggga agacaatagc aggcatgaga tctcacgtgc ggaccgagcg
     3961 gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac
     4021 tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag
     4081 cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct
     4141 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
     4201 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta
     4261 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     4321 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     4381 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     4441 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     4501 acaacactca actctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     4561 gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     4621 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     4681 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     4741 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     4801 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     4861 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     4921 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     4981 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     5041 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     5101 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     5161 ctggatctca acagcggtaa gatc
//
