LOCUS       sequence-466321-        4759 bp    DNA     circular SYN 15-APR-2025
DEFINITION  AiE2463m 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 4759)
  AUTHORS   Ben-Simon Y, Hooper M, Narayan S, Daigle T, Dwivedi D, Way SW, Oster
            A, Stafford DA, Mich JK, Taormina MJ, Martinez RA, Opitz-Araya X, 
            Roth JR, Allen S, Ayala A, Bakken TE, Barcelli T, Barta S, Bendrick 
            J, Bertagnolli D, Bowlus J, Boyer G, Brouner K, Casian B, Casper T, 
            Chakka AB, Chakrabarty R, Chance RK, Chavan S, Departee M, Donadio 
            N, Dotson N, Egdorf T, Gabitto M, Garcia J, Gary A, Gasperini M, 
            Goldy J, Gore BB, Graybuck L, Greisman N, Haeseleer F, Halterman C, 
            Helback O, Hockemeyer D, Huang C, Huff S, Hunker A, Johansen N, 
            Juneau Z, Kalmbach B, Khem S, Kussick E, Kutsal R, Larsen R, Lee C, 
            Lee AY, Leibly M, Lenz GH, Liang E, Lusk N, Malone J, Mollenkopf T, 
            Morin E, Newman D, Ng L, Ngo K, Omstead V, Oyama A, Pham T, Pom CA, 
            Potekhina L, Ransford S, Rette D, Rimorin C, Rocha D, Ruiz A, 
            Sanchez REA, Sedeno-Cortes A, Sevigny JP, Shapovalova N, Shulga L, 
            Sigler AR, Siverts LA, Somasundaram S, Stewart K, Szelenyi E, Tieu 
            M, Trader C, van Velthoven CTJ, Walker M, 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 J, Tasic B
  TITLE     A suite of enhancer AAVs and transgenic mouse lines for genetic 
            access to cortical cell types.
  JOURNAL   bioRxiv [Preprint]. 2024 Sep 26:2024.06.10.597244. doi: 
            10.1101/2024.06.10.597244.
  PUBMED    38915722
REFERENCE   2  (bases 1 to 4759)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2024 Sep 26:2024.06.10.597244. doi: 
            10.1101/2024.06.10.597244."
COMMENT     Sequence Label: AiP1881 - pAAV-AiE2463m-minBG-SYFP2-WPRE3-BGHpA
FEATURES             Location/Qualifiers
     source          1..4759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(428..1219,1220..1288))
                     /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     1051..1070
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1289..1393)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1461..1479
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1517..1539)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1639..1658
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(1675..2130)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1812..1833)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2024..2043
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2205..2345
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2216..2345
                     /label=AAV2 ITR
     polyA_signal    2463..2574
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2761..2781
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(2874..3584,3585..3587,3588..3590))
                     /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(2916..2937)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3265..3284
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3524..3545
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3587..3596
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     repeat_region   4237..4366
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(join(4428..4759,1..257))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4508..4527)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
       61 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
      121 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
      181 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
      241 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
      301 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
      361 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
      421 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
      481 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
      541 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
      601 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
      661 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
      721 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
      781 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
      841 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
      901 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
      961 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     1021 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     1081 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     1141 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     1201 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     1261 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     1321 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     1381 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta
     1441 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg
     1501 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     1561 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     1621 ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac cataaaattg
     1681 taaacgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     1741 accaatagac cgaaatcggc aaaatccctt ataaatcaaa agaatagccc gagatagagt
     1801 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     1861 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca cccaaatcaa
     1921 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     1981 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     2041 gagcgggcgc taaggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     2101 ccgcgcttaa tgcgccgcta cagggcgcgt actatggttg ctttgacgta tgcggtgtga
     2161 aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgcccctgca ggcagctgcg
     2221 cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg
     2281 cccggcctca gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg
     2341 ttcctgcggc cgctcggtcc gcacgtgaga tctcatgcct gctattgtct tcccaatcct
     2401 cccccttgct gtcctgcccc accccacccc ccagaataga atgacaccta ctcagacaat
     2461 gcgatgcaat ttcctcattt tattaggaaa ggacagtggg agtggcacct tccagggtca
     2521 aggaaggcac gggggagggg caaacaacag atggctggca actagaaggc acagtcgaag
     2581 atctctcgag ccacggaatt gtcagtgccc aacagccgag cccctgtcca gcagcgggca
     2641 aggcaggcgg cgatgagttc cgccgtggca agaactaacc aggatttata caaggaggag
     2701 aaaatgaaag ccatacggga agcaatagca tgatacaaag gcattaaagc agcgtatcca
     2761 catagcgtaa aaggagcaac atagttaaga ataccagtca atctttcaca aattttgtaa
     2821 tccagaggtt gattatgata tcgaattcgc ggccgcggcg cgccgtcgac ttacttgtac
     2881 agctcgtcca tgccgagagt gatcccggcg gcggtcacga actccagcag gaccatgtga
     2941 tcgcgcttct cgttggggtc tttgctcagc ttggactggt agctcaggta gtggttgtcg
     3001 ggcagcagca cggggccgtc gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc
     3061 acgccgccgt cctcgatgtt gtggcggatc ttgaagttgg ccttgatgcc gttcttctgc
     3121 ttgtcggcgg tgatatagac gttgtggctg ttgtagttgt actccagctt gtgccccagg
     3181 atgttgccgt cctccttgaa gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg
     3241 ccctcgaact tcacctcggc gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg
     3301 cgctcctgga cgtagccttc gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg
     3361 tcggggtagc gggcgaagca ctgcacgccg tagcccaggg tggtcacgag ggtgggccag
     3421 ggcacgggca gcttgccggt ggtgcagatc agcttcaggg tcagcttgcc gtaggtggca
     3481 tcgccctcgc cctcgccgga cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg
     3541 accaggatgg gcaccacccc ggtgaacagc tcctcgccct tgctcaccat ggtggcgacc
     3601 ggtagcgcta gcttcgaaag atctggatcc cagaagcaaa tgtaagcaat agatggctct
     3661 gccctgactt ttatgcccag cccgagctcc ggtcacagtt cttttctcta aagttcttgt
     3721 tcttattaat gtggccatta tgctcattga tttcttcagc ctctttgttc caaggtactg
     3781 cttcaaattt ggagtcaaat gagcattcct tggaacatgt acttgtacca aacagcctta
     3841 gccagaagaa ggtgggagtg ggtgtatttt actgaaatta gagtattctt cataaacgag
     3901 agaatatagt ggagagaaaa taagtgcaga gttacagaaa tgtttcttat tctgccaaat
     3961 tcatttatcc tcaagcatcc tctagatggt attttgatgt gttattggta aatcattcta
     4021 atgtcttgtc ttcattctag gtggatattt attttgattc aacctgacag ctagcaaaca
     4081 ttctgttata aaaagccagg ttttgaaaag cagggatgtt accaagggga tgagaagaga
     4141 agctctcaga tcatgccaac tcaaactgtg aagtaggaaa aaaatgatat attttgtctc
     4201 tgtctctctg tccctctgcc tctgtcttgc ggccgcagga acccctagtg atggagttgg
     4261 ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac
     4321 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggacat gtgagcaaaa
     4381 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     4441 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     4501 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     4561 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     4621 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     4681 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     4741 tccaacccgg taagacacg
//
