LOCUS       sequence-447005-        5918 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0441h_3xC2 is an enhancer sequence, designed to drive Cre 
            dependent AAV-mediated transgene expression in striatal MSNs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5918)
  AUTHORS   Hunker AC, Wirthlin ME, Gill G, Johansen NJ, Hooper M, Omstead V, 
            Vargas S, Lerma MN, Taskin N, Weed N, Laird WD, Bishaw YM, Bendrick 
            JL, Gore BB, Ben-Simon Y, Opitz-Araya X, Martinez RA, Way SW, 
            Thyagarajan B, Otto S, Sanchez REA, Alexander JR, Amaya A, Amster A,
            Arbuckle J, Ayala A, Baker PM, Barcelli T, Barta S, Bertagnolli D, 
            Bielstein C, Bishwakarma P, Bowlus J, Boyer G, Brouner K, Casian B, 
            Casper T, Chakka AB, Chakrabarty R, Chong P, Clark M, Colbert K, 
            Daniel S, Dawe T, Departee M, DiValentin P, Donadio NP, Dotson NI, 
            Dwivedi D, Egdorf T, Fliss T, Gary A, Goldy J, Grasso C, Groce EL, 
            Gudsnuk K, Han W, Haradon Z, Hastings S, Helback O, Ho WV, Huang C, 
            Johnson T, Jones DL, Juneau Z, Kenney J, Leibly M, Li S, Liang E, 
            Loeffler H, Lusk NA, Madigan Z, Malloy J, Malone J, McCue R, Melchor
            J, Mich JK, Moosman S, Morin E, Naidoo R, Newman D, Ngo K, Nguyen K,
            Oster AL, Ouellette B, Oyama AA, Pena N, Pham T, Phillips E, Pom C, 
            Potekhina L, Ransford S, Ray PL, Reding M, Rette DF, Reynoldson C, 
            Rimorin C, Sigler AR, Rocha DB, Ronellenfitch K, Ruiz A, Sawyer L, 
            Sevigny JP, Shapovalova NV, Shepard N, Shulga L, Soliman S, Staats 
            B, Taormina MJ, Tieu M, Wang Y, Wilkes J, Wood T, Zhou T, Williford 
            A, Dee N, Mollenkopf T, Ng L, Esposito L, Kalmbach BE, Yao S, Ariza 
            J, Collman F, Mufti S, Smith K, Waters J, Ersing I, Patrick M, Zeng 
            H, Lein ES, Kojima Y, Horwitz G, Owen SF, Levi BP, Daigle TL, Tasic 
            B, Bakken TE, Ting JT
  TITLE     Enhancer AAV toolbox for accessing and perturbing striatal cell 
            types and circuits.
  JOURNAL   Neuron. 2025 May 21;113(10):1507-1524.e17. doi: 
            10.1016/j.neuron.2025.04.035.
  PUBMED    40403704
REFERENCE   2  (bases 1 to 5918)
  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"; doi: 
            "10.1016/j.neuron.2025.04"; journalName: "Neuron"; date: 
            "2025-05-21- 21"; volume: "113"; issue: "10"; pages: "1507-1524.e17"
COMMENT     Sequence Label: AiP15357 -
            pAAV-AiE0441h_3xC2-minBG-DIO-CoChR-...
FEATURES             Location/Qualifiers
     source          1..5918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     protein_bind    complement(825..858)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(909..942)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(951..1664,1665..1667))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             complement(join(954..1664,1665..1667))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(996..1017)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1345..1364
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1604..1625
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             complement(1686..2549)
                     /codon_start=1
                     /product="high-photocurrent channelrhodopsin from 
                     Chloromonas oogama (Shemesh et al., 2017)"
                     /label=CoChR
                     /translation="MLGNGSAIVPIDQCFCLAWTDSLGSDTEQLVANILQWFAFGFSIL
                     ILMFYAYQTWRATCGWEEVYVCCVELTKVIIEFFHEFDDPSMLYLANGHRVQWLRYAEW
                     LLTCPVILIHLSNLTGLKDDYSKRTMRLLVSDVGTIVWGATSAMSTGYVKVIFFVLGCI
                     YGANTFFHAAKVYIESYHVVPKGRPRTVVRIMAWLFFLSWGMFPVLFVVGPEGFDAISV
                     YGSTIGHTIIDLMSKNCWGLLGHYLRVLIHQHIIIYGDIRKKTKINVAGEEMEVETMVD
                     QEDEETV"
     protein_bind    2560..2593
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    2644..2677
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(2745..2765)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2952..3063
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3181..3321
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3181..3310
                     /label=AAV2 ITR
     rep_origin      3396..3851
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3483..3502)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3693..3714
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3868..3887)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3987..4009
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4047..4065)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4133..4237
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4238..4306,4307..5098)
                     /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(4456..4475)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5269..5857
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5758..5777
                     /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 gcgtcccaca cagcccccac cacctggccg cccacctgaa
      181 aaacttttcc ccaagttctt gagaagtcgc gtaggatcca ggagagaaaa atcaaagtaa
      241 taaaacctca tttaattcct aacggagggc agagctgcga gctgcaatta tcttaatgct
      301 gagctatttt tactctctct ccatctctgt cttctgcctt tctcccacac agcccccacc
      361 acctggccgc ccacctgaaa aacttttccc caagttcttg agaagtcgcg taggatccag
      421 gagagaaaaa tcaaagtaat aaaacctcat ttaattccta acggagggca gagctgcgag
      481 ctgcaattat cttaatgctg agctattttt actctctctc catctctgtc ttctgccttt
      541 ctcccacaca gcccccacca cctggccgcc cacctgaaaa acttttcccc aagttcttga
      601 gaagtcgcgt aggatccagg agagaaaaat caaagtaata aaacctcatt taattcctaa
      661 cggagggcag agctgcgagc tgcaattatc ttaatgctga gctattttta ctctctctcc
      721 atctctgtct tctgcctttc tgagctcggg ctgggcataa aagtcagggc agagccatct
      781 attgcttaca tttgcttctg ggatcctcta gagtcgactc cggaataact tcgtatagga
      841 tactttatac gaagttatgc agaatggtag ctggattgta gctgctatta gcaatatgaa
      901 acctcttaat aacttcgtat agcatacatt atacgaagtt atggcgcgcc ttacttgtac
      961 agctcgtcca tgccgagagt gatcccggcg gcggtcacga actccagcag gaccatgtga
     1021 tcgcgcttct cgttggggtc tttgctcagg gcggactggg tgctcaggta gtggttgtcg
     1081 ggcagcagca cggggccgtc gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc
     1141 acgctgccgt cctcgatgtt gtggcggatc ttgaagttca ccttgatgcc gttcttctgc
     1201 ttgtcggcca tgatatagac gttgtggctg ttgtagttgt actccagctt gtgccccagg
     1261 atgttgccgt cctccttgaa gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg
     1321 ccctcgaact tcacctcggc gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg
     1381 cgctcctgga cgtagccttc gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg
     1441 tcggggtagc ggctgaagca ctgcacgccg taggtcaggg tggtcacgag ggtgggccag
     1501 ggcacgggca gcttgccggt ggtgcaaatg aacttcaggg tcagcttgcc gtaggtggca
     1561 tcgccctcgc cctcgccgga cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg
     1621 accaggatgg gcaccacccc ggtgaacagc tcctcgccct tgctcactgc tactaccggt
     1681 gccgccactg tctcctcgtc ctcctggtcc accatggtct ccacttccat ctcctcgccg
     1741 gccacgttga tcttggtctt cttccggatg tcgccgtaga tgatgatgtg ctggtggatc
     1801 agcactctca gatagtgtcc cagcaggccc caacaattct tgctcatgag gtcgatgatg
     1861 gtgtggccaa tggtagagcc gtacacgctg atggcgtcga atccttctgg tcccacgacg
     1921 aacagcacgg ggaacatgcc ccaagacagg aagaacagcc aagccatgat ccgcacgacg
     1981 gttctaggtc tgccctttgg caccacgtgg tagctctcga tatacacctt ggcggcgtgg
     2041 aagaaggtgt tggcgccgta gatgcaaccc agcacgaaga agatgacctt gacgtagcct
     2101 gtgctcatgg cgcttgtagc tccccacacg atggttccca cgtctgacac cagcagcctc
     2161 atggtccgct tgctgtagtc gtccttcagg ccggtcaggt tgctcaggtg gatcaggatg
     2221 acgggacaag tcagcagcca ctctgcgtat ctcagccact ggactcggtg tccgttagcc
     2281 aggtacagca tgctggggtc gtcgaactcg tggaagaact cgatgatgac cttggtcagc
     2341 tcgacgcaac agacgtagac ctcctcccaa ccgcaagtgg ctctccaagt ctggtaggcg
     2401 tagaacatca ggatcaggat gctgaagccg aaggcgaacc actggaggat gttggccacc
     2461 agctgctctg tatcgcttcc caggctgtcg gtccaagcca ggcaaaagca ctggtcgata
     2521 ggcacaatgg cgctgccgtt tcccagcatg gtggctagca taacttcgta taaagtatcc
     2581 tatacgaagt tatttgcctt aacccagaaa ttatcactgt tattctttag aatggtgcaa
     2641 agaataactt cgtataatgt atgctatacg aagttatgaa ttcgatatca taatcaacct
     2701 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
     2761 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
     2821 attttctcct ccttgtataa atcctggtta gttcttgcca cggcggaact catcgccgcc
     2881 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggctcga
     2941 gagatcttcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     3001 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     3061 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     3121 gggggaggat tgggaagaca atagcaggca tgagatctca cgtgcggacc gagcggccgc
     3181 aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg
     3241 ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc
     3301 gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt tctccttacg catctgtgcg
     3361 gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag
     3421 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc
     3481 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     3541 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     3601 aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     3661 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     3721 actcaactct atctcgggct attcttttga tttataaggg attttgccga tttcggtcta
     3781 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     3841 gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     3901 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     3961 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     4021 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     4081 catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac
     4141 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     4201 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     4261 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     4321 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     4381 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     4441 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     4501 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     4561 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     4621 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     4681 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     4741 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     4801 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     4861 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     4921 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     4981 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     5041 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     5101 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     5161 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     5221 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     5281 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     5341 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     5401 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     5461 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     5521 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     5581 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     5641 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     5701 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     5761 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     5821 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     5881 acggttcctg gccttttgct ggccttttgc tcacatgt
//
