LOCUS       sequence-466230-        6607 bp    DNA     circular SYN 15-APR-2025
DEFINITION  AiE2096m 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 6607)
  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 6607)
  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: AiP1265 - pAAV-AiE2096m-minBG-FlpO-WPRE-HGHpA
FEATURES             Location/Qualifiers
     source          1..6607
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     253..272
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   414..543
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      1818..1827
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1824..1829,1830..1850,1851..3122)
                     /codon_start=1
                     /product="nuclear-targeted site-specific recombinase"
                     /label=FLPo
                     /note="FLPo is a mouse codon-optimized fusion between the 
                     SV40 nuclear localization signal and a thermostable version
                     of the Saccharomyces cerevisiae site-specific recombinase 
                     FLP.  Recombination occurs at FRT sequences."
                     /translation="MAPKKKRKVMSQFDILCKTPPKVLVRQFVERFERPSGEKIASCAA
                     ELTYLCWMITHNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKK
                     LIPAWEFTIIPYNGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESI
                     WEITEKILNSFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGV
                     IIQCLVTETKTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEY
                     QLLKDNLVRSYNKALKKNAPYPIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWS
                     DKRASAVARTTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQL
                     KGSAEGSIRYPAWNGIISQEVLDYLSSYINRRI"
     misc_feature    3147..3735
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3200..3220)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3618..3629)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3767..4243
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3966..3985)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4283..4423
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4283..4412
                     /label=AAV2 ITR
     rep_origin      4498..4953
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4585..4604)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4795..4816
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4970..4989)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5089..5111
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5149..5167)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5235..5339
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5340..5408,5409..6200)
                     /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(5558..5577)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(6371..6607,1..352)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
       61 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
      121 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
      181 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
      241 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
      301 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
      361 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgtcctgcag
      421 gcagctgcgc gctcgctcgc tcactgaggc cgcccgggcg tcgggcgacc tttggtcgcc
      481 cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc actaggggtt
      541 cctgcggccg cacgcgtttt catcaggatc tgctataggg agccttggag aaggccttct
      601 gctatgagct gctgtctaat ctcccttaga aggcctctct gtactcccaa ccatggcttc
      661 cagcaagcca agctcaagcc tgctgctgtc aagccaagaa ctccccccat ggggccagct
      721 ggagctcccc tcttttttcc ctcagcccct ggctagatcc agatcatggg cccacattcc
      781 gttcttggct cttctgtggc atccagtggc gcctggatgt ccaagagcct gataaccaga
      841 ctgccctggc cttcttgcag gcctgggaac tcctgagcca gttctgggtg tcccacacct
      901 cagactgttc ttttccaccc cagggacagg gtcccagggc ttccctacag tccaatacat
      961 gcctggcaac agcatggggt aagcacagtc aacccccgtg tccctcgtcc ctgagtggtg
     1021 gagccctgtg gcagtgcaga caggggacca ataaccctac acatgagtct ctattctatg
     1081 tatctattgg ggaggaggat ttggattccg tgtattagga attggctgga ctccttgcat
     1141 tgtaagtagt ctggaactgg tgtgttctag aagacagaat atcccattat aatttgaata
     1201 ttggcagatt ctgctcttgg cctaggatat aggacatgaa gttctgtagt acagctgttc
     1261 tcatcacttt aactttagag actaggattg agatccttgg gtctttatga aaacatacag
     1321 gtggttataa attgagaatg acatgttctt tagatcagta tgaacagata agatgactgc
     1381 gtcctcaagg gacaattatc ccaacaccaa ggagaaaaac tcatctagaa cagaagttat
     1441 taaaccagca gagttgtaac atatggtgtc atttagctgt ttcttttgaa aagaagtttg
     1501 aagtccagca ggcccacaga tacaggggtt ttcctctgtg tgctagggta ggtcctttgg
     1561 tggtggggtc atctatttgt gacttgatca tacctattga tttctagtag tctgacagcc
     1621 atgagtatgc tcaaagccac ttggtaaagc ctcttccatt gaggacccaa ctgttttttg
     1681 gatcctttcc tcccaggttt taagaaaggc caactcccca gggtctacaa tattcagagc
     1741 cacacccctt aaggagctcg ggctgggcat aaaagtcagg gcagagccat ctattgctta
     1801 catttgcttc tggcgtggcc accatggctc ctaagaagaa gaggaaggtg atgagccagt
     1861 tcgacatcct gtgcaagacc ccccccaagg tgctggtgcg gcagttcgtg gagagattcg
     1921 agaggcccag cggcgagaag atcgccagct gtgccgccga gctgacctac ctgtgctgga
     1981 tgatcaccca caacggcacc gccatcaaga gggccacctt catgagctac aacaccatca
     2041 tcagcaacag cctgagcttc gacatcgtga acaagagcct gcagttcaag tacaagaccc
     2101 agaaggccac catcctggag gccagcctga agaagctgat ccccgcctgg gagttcacca
     2161 tcatccctta caacggccag aagcaccaga gcgacatcac cgacatcgtg tccagcctgc
     2221 agctgcagtt cgagagcagc gaggaggccg acaagggcaa cagccacagc aagaagatgc
     2281 tgaaggccct gctgtccgag ggcgagagca tctgggagat caccgagaag atcctgaaca
     2341 gcttcgagta caccagcagg ttcaccaaga ccaagaccct gtaccagttc ctgttcctgg
     2401 ccacattcat caactgcggc aggttcagcg acatcaagaa cgtggacccc aagagcttca
     2461 agctggtgca gaacaagtac ctgggcgtga tcattcagtg cctggtgacc gagaccaaga
     2521 caagcgtgtc caggcacatc tactttttca gcgccagagg caggatcgac cccctggtgt
     2581 acctggacga gttcctgagg aacagcgagc ccgtgctgaa gagagtgaac aggaccggca
     2641 acagcagcag caacaagcag gagtaccagc tgctgaagga caacctggtg cgcagctaca
     2701 acaaggccct gaagaagaac gccccctacc ccatcttcgc tatcaagaac ggccctaaga
     2761 gccacatcgg caggcacctg atgaccagct ttctgagcat gaagggcctg accgagctga
     2821 caaacgtggt gggcaactgg agcgacaaga gggcctccgc cgtggccagg accacctaca
     2881 cccaccagat caccgccatc cccgaccact acttcgccct ggtgtccagg tactacgcct
     2941 acgaccccat cagcaaggag atgatcgccc tgaaggacga gaccaacccc atcgaggagt
     3001 ggcagcacat cgagcagctg aagggcagcg ccgagggcag catcagatac cccgcctgga
     3061 acggcatcat cagccaggag gtgctggact acctgagcag ctacatcaac aggcggatct
     3121 gagaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat
     3181 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     3241 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     3301 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     3361 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     3421 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     3481 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     3541 aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg cgcgggacgt
     3601 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     3661 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     3721 gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt ggcatccctg
     3781 tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc ccaccagcct
     3841 tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct ataatattat
     3901 ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct gtagggcctg
     3961 cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca ctgcaatctc
     4021 cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg gattccaggc
     4081 atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt tcaccatatt
     4141 ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc ctcccaaatt
     4201 gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt tgtaggtaac
     4261 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     4321 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     4381 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     4441 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     4501 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4561 cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4621 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4681 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     4741 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4801 tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt gatttataag
     4861 ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4921 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     4981 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     5041 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     5101 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     5161 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     5221 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     5281 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     5341 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5401 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     5461 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5521 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5581 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     5641 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     5701 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     5761 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     5821 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     5881 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     5941 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     6001 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     6061 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     6121 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     6181 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     6241 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     6301 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6361 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6421 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6481 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     6541 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     6601 cagtggc
//
