LOCUS       sequence-474114-        5400 bp    DNA     circular SYN 30-MAY-2025
DEFINITION  AiE0546h 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 5400)
  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 5400)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 30, 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: AiP13651 -
            pAAV-AiE0546h-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5400
                     /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      866..875
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(872..874,875..877,878..1588)
                     /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(1178..1197)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1525..1546
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1598..1654
                     /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             1667..1732
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1763..2140
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(2227..2247)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2434..2545
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2663..2803
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2663..2792
                     /label=AAV2 ITR
     rep_origin      2878..3333
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2965..2984)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3175..3196
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3350..3369)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3469..3491
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3529..3547)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3615..3719
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3720..3788,3789..4580)
                     /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(3938..3957)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4751..5339
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5240..5259
                     /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 gcgtttaagg ctgcagtgag ccacgatcat gccactgcac
      181 tccagcctgg gtgacagagc gagactctgt ctccaaaaat ataataaaat aaaataaaat
      241 aaaaataaaa ataaaggatt ctgcaaatgt cagcctactc ctctgcccaa aatatataat
      301 aacaaagaaa acaaagcaaa gatgtcatag ctatggtttt gttgctatga gaatactctt
      361 gtcaaggctg gaggatggca caaacatact gtatttaacc acttgtcagt ttcacttgca
      421 attcacacac gtggcacgtg ggcctctgtg ctcttgtcct gggtcagcca atgtcagggg
      481 cgggagcgag ggtacgaatg cttcgagaat ggacccggca agttgaatgc ttccgagagg
      541 ttaggatgag gccgccaggg tgagcacgtg cggatgatgg gtggggcggt gagttgttgc
      601 tgggtagtgc agaagaaact gttctggggt aatgcacgtg ggtgcaggtg ggaggggact
      661 ggagaagggt gcaggtggga atgagatcat aggggaggat acagtgctgc aggaaatata
      721 ttttttaaat ttttaaattt tatttttatt tttagatgga gtctctgttg ccgagctcgg
      781 gctgggcata aaagtcaggg cagagccatc tattgcttac atttgcttct gggatccaga
      841 tctttcgaag ctagcgctac cggtcgccac catggtctcc aagggcgagg agctgttcac
      901 cggggtggtg cccatcctgg tcgagctgga cggcgatgtc aacggccaca agttcagcgt
      961 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagc tgatctgcac
     1021 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgggct acggcgtgca
     1081 gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     1141 cgaaggctac gtccaggagc gcaccatctt cttcaaagac gacggcaact acaagacccg
     1201 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     1261 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     1321 cgtctatatc accgccgaca agcagaagaa cggcatcaag gccaacttca agatccgcca
     1381 caacatcgag gacggcggcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     1441 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc taccagtcca agctgagcaa
     1501 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     1561 cactctcggc atggacgagc tgtacaaggg ctctggtgct accaacttct cactgttgaa
     1621 acaggcaggg gatgtagagg agaatccagg gcctggtgct agtggagact acaaagacca
     1681 tgacggagat tataaagatc atgacatcga ttacaaggat gacgatgaca agtccggact
     1741 cagatctgga ggctccggag gcccagagcc agcgaagtct gctcccgccc cgaaaaaggg
     1801 ctccaagaag gcggtgacta aggcgcagaa gaaaggcggc aagaagcgca agcgcagccg
     1861 caaggagagc tattccatct atgtgtacaa ggttctgaag caggtccacc ctgacaccgg
     1921 catttcgtcc aaggccatgg gcatcatgaa ttcgtttgtg aacgacattt tcgagcgcat
     1981 cgcaggagag gcttcccgcc tggcgcatta caacaagcgc tcgaccatca cctcccggga
     2041 gatccagacg gccgtgcgcc tgctgctgcc tggggagttg gccaagcacg ccgtgtccga
     2101 gggtactaag gccatcacca agtacaccag cgctaagtaa tgaggcgcgc cgcggccgcg
     2161 aattcgatat cataatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     2221 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     2281 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tagttcttgc
     2341 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     2401 cactgacaat tccgtggctc gagagatctt cgactgtgcc ttctagttgc cagccatctg
     2461 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     2521 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     2581 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgagatct
     2641 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     2701 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     2761 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     2821 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     2881 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     2941 cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     3001 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     3061 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     3121 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     3181 tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt gatttataag
     3241 ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     3301 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     3361 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     3421 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     3481 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     3541 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     3601 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     3661 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     3721 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     3781 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     3841 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     3901 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     3961 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     4021 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     4081 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     4141 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     4201 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     4261 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     4321 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     4381 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     4441 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     4501 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     4561 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     4621 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     4681 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     4741 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     4801 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     4861 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     4921 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     4981 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     5041 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     5101 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     5161 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     5221 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     5281 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     5341 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
