LOCUS       sequence-460627-        5449 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0324m 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 5449)
  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 5449)
  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: AiP14536 -
            pAAV-AiE0324m-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5449
                     /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      802..811
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(808..810,811..813,814..1524)
                     /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(1114..1133)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1461..1482
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1534..1590
                     /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             1603..1668
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1699..2076
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(2163..2183)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2370..2481
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2599..2739
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2599..2728
                     /label=AAV2 ITR
     misc_feature    2746..2773
                     /gene="telR"
                     /label=telR
                     /note="telomeric sequence from coliphage N15"
     misc_feature    2774..2801
                     /gene="telL"
                     /label=telL
                     /note="telomeric sequence from coliphage N15"
     rep_origin      2870..3325
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2957..2976)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3167..3188
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3342..3361)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3461..3483
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3521..3539)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3607..3711
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3712..3780,3781..4572)
                     /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(3930..3949)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4743..5331
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5232..5251
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    5388..5415
                     /gene="telR"
                     /label=telR
                     /note="telomeric sequence from coliphage N15"
     misc_feature    5416..5443
                     /gene="telL"
                     /label=telL
                     /note="telomeric sequence from coliphage N15"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtcctgcc atctccgact cacagcattc tgctgctcct
      181 agctctaaga ccggaggggt gtggtgaacg agaggattac atctgtcaga ggcaagaatc
      241 atgcattgga gggcagagag gtccagggtt tttccctgag cagggctggg agtgggctct
      301 gatgcggtaa gcatttgatg tggtgtgtca ctgccttatt ccttgcagag agtgggtgtt
      361 tgctgtcagg agcaggagcc agaggaaggg gcctgatgca cactcgttgg ctgtttcctt
      421 ggacagccat ctgcccacag cctataattc agaggcccag ctgcactggg tgcttgaccg
      481 atctgattgt gaagagggga gattaccaag tgcactgcag tgattggcag gtgccccagg
      541 gcccagctgc cctggtagca gagtggactc caagggacac tgttctatct ctgaatgccc
      601 acctcccaat tattcaggcc tcccagaatc ccacagtcac ctgcaatcct cttgatcctt
      661 agaacccctg cttcccccag agtgggtgaa tgtgcaaaca catcccgtga gctcgggctg
      721 ggcataaaag tcagggcaga gccatctatt gcttacattt gcttctggga tccagatctt
      781 tcgaagctag cgctaccggt cgccaccatg gtctccaagg gcgaggagct gttcaccggg
      841 gtggtgccca tcctggtcga gctggacggc gatgtcaacg gccacaagtt cagcgtgtcc
      901 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagctgat ctgcaccacc
      961 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgggctacgg cgtgcagtgc
     1021 ttcgcccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
     1081 ggctacgtcc aggagcgcac catcttcttc aaagacgacg gcaactacaa gacccgcgcc
     1141 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     1201 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     1261 tatatcaccg ccgacaagca gaagaacggc atcaaggcca acttcaagat ccgccacaac
     1321 atcgaggacg gcggcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     1381 ggccccgtgc tgctgcccga caaccactac ctgagctacc agtccaagct gagcaaagac
     1441 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     1501 ctcggcatgg acgagctgta caagggctct ggtgctacca acttctcact gttgaaacag
     1561 gcaggggatg tagaggagaa tccagggcct ggtgctagtg gagactacaa agaccatgac
     1621 ggagattata aagatcatga catcgattac aaggatgacg atgacaagtc cggactcaga
     1681 tctggaggct ccggaggccc agagccagcg aagtctgctc ccgccccgaa aaagggctcc
     1741 aagaaggcgg tgactaaggc gcagaagaaa ggcggcaaga agcgcaagcg cagccgcaag
     1801 gagagctatt ccatctatgt gtataaggtt ctgaagcagg tccaccctga caccggcatt
     1861 tcgtccaagg ccatgggcat catgaactcg tttgtgaacg acattttcga gcgcatcgca
     1921 ggtgaggctt cccgcctggc gcattacaac aagcgctcga ccatcacctc cagggagatc
     1981 cagacggccg tgcgcctgct gctgcctggg gagttggcca agcacgccgt gtccgagggt
     2041 actaaggcca tcaccaagta caccagcgct aagtaatgag gcgcgccgcg gccgcgaatt
     2101 cgatatcata atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     2161 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     2221 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttagt tcttgccacg
     2281 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
     2341 gacaattccg tggctcgaga gatcttcgac tgtgccttct agttgccagc catctgttgt
     2401 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     2461 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     2521 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg agatctcacg
     2581 tgcggaccga gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg
     2641 ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg
     2701 cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctatca gcacacaatt
     2761 gcccattata cgcgcgtata atggactatt gtgtgctgat atgatgcggt attttctcct
     2821 tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta cgcgccctgt
     2881 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     2941 agcgccttag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     3001 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     3061 cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga
     3121 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     3181 caaactggaa caacactcaa ctctatctcg ggctattctt ttgatttata agggattttg
     3241 ccgatttcgg tctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     3301 aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg ctctgatgcc
     3361 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     3421 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     3481 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt
     3541 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
     3601 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     3661 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     3721 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     3781 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     3841 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     3901 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     3961 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     4021 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     4081 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     4141 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     4201 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     4261 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     4321 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     4381 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     4441 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     4501 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     4561 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     4621 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     4681 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     4741 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     4801 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     4861 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     4921 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     4981 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     5041 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     5101 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     5161 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     5221 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     5281 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     5341 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcatat cagcacacaa
     5401 ttgcccatta tacgcgcgta taatggacta ttgtgtgctg ataacatgt
//
