LOCUS       sequence-459520-        6311 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AiE0873m_3xC2 is an enhancer sequence, designed to drive 
            AAV-mediated transgene expression in striatal cholinergic 
            interneurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6311)
  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 6311)
  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: AiP15381:
            pAAV-AiE0873m_3xC2-minBG-SYFP2-P2A-m...
FEATURES             Location/Qualifiers
     source          1..6311
                     /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      1147..1156
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1153..1155,1156..1158,1159..1869)
                     /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(1198..1219)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1459..1478)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1806..1827
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1879..1935
                     /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             1951..2646
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(2084..2102)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2352..2371
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2677..3054
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(3138..3158)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    3345..3456
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3574..3714
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3574..3703
                     /label=AAV2 ITR
     rep_origin      3789..4244
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3876..3895)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4086..4107
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4261..4280)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4380..4402
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4440..4458)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4526..4630
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4631..4699,4700..5491)
                     /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(4849..4868)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5662..6250
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6151..6170
                     /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 gcgtcatgga tttctccaaa gctttgtagg gcagaactct
      181 gaatgtttat ttctctgatg aatccttaat gaaagtggcc agccatcctg gggagaacaa
      241 gccttagttc ccactgagaa ggatagcaag cagcataaga gtagctgatg tcaaaggcta
      301 ttaatgcaaa tgaaaattat aattcctcat caagatctgc tcccagacag gctaatcatt
      361 gtgccacctg tttacagaaa cacactgcta atcagtcagt aaaatctttt aagtttctgt
      421 gcttctctgt gtacgatttt tagcctccat ggatttctcc aaagctttgt agggcagaac
      481 tctgaatgtt tatttctctg atgaatcctt aatgaaagtg gccagccatc ctggggagaa
      541 caagccttag ttcccactga gaaggatagc aagcagcata agagtagctg atgtcaaagg
      601 ctattaatgc aaatgaaaat tataattcct catcaagatc tgctcccaga caggctaatc
      661 attgtgccac ctgtttacag aaacacactg ctaatcagtc agtaaaatct tttaagtttc
      721 tgtgcttctc tgtgtacgat ttttagcctc catggatttc tccaaagctt tgtagggcag
      781 aactctgaat gtttatttct ctgatgaatc cttaatgaaa gtggccagcc atcctgggga
      841 gaacaagcct tagttcccac tgagaaggat agcaagcagc ataagagtag ctgatgtcaa
      901 aggctattaa tgcaaatgaa aattataatt cctcatcaag atctgctccc agacaggcta
      961 atcattgtgc cacctgttta cagaaacaca ctgctaatca gtcagtaaaa tcttttaagt
     1021 ttctgtgctt ctctgtgtac gatttttagc ctcgagctcg ggctgggcat aaaagtcagg
     1081 gcagagccat ctattgctta catttgcttc tgggatccag atctttcgaa gctagcgcta
     1141 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     1201 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     1261 gatgccacct acggcaagct gaccctgaag ctgatctgca ccaccggcaa gctgcccgtg
     1321 ccctggccca ccctcgtgac caccctgggc tacggcgtgc agtgcttcgc ccgctacccc
     1381 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     1441 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     1501 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1561 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat caccgccgac
     1621 aagcagaaga acggcatcaa ggccaacttc aagatccgcc acaacatcga ggacggcggc
     1681 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1741 cccgacaacc actacctgag ctaccagtcc aagctgagca aagaccccaa cgagaagcgc
     1801 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1861 ctgtacaagg gaagcggagc cactaacttc tccctgttga aacaagcagg ggatgtcgaa
     1921 gagaatcccg ggccagcacc gggctccacc atggtgagca agggcgaggc agtgatcaag
     1981 gagttcatgc ggttcaaggt gcacatggag ggctccatga acggccacga gttcgagatc
     2041 gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc
     2101 aagggtggcc ccctgccctt ctcctgggac atcctgtccc ctcagttcat gtacggctcc
     2161 agggccttca ccaagcaccc cgccgacatc cccgactact ataagcagtc cttccccgag
     2221 ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgccgtgac cgtgacccag
     2281 gacacctccc tggaggacgg caccctgatc tacaaggtga agctccgcgg caccaacttc
     2341 cctcctgacg gccccgtaat gcagaagaag acaatgggct gggaagcgtc caccgagcgg
     2401 ttgtaccccg aggacggcgt gctgaagggc gacattaaga tggccctgcg cctgaaggac
     2461 ggcggccgct acctggcgga cttcaagacc acctacaagg ccaagaagcc cgtgcagatg
     2521 cccggcgcct acaacgtcga ccgcaagttg gacatcacct cccacaacga ggactacacc
     2581 gtggtggaac agtacgaacg ctccgagggc cgccactcca ccggcggcat ggacgagctg
     2641 tataagtccg gactcagatc tggaggctcc ggaggcccag agccagcgaa gtctgctccc
     2701 gccccgaaaa agggctccaa gaaggcggtg actaaggcgc agaagaaagg cggcaagaag
     2761 cgcaagcgca gccgcaagga gagctattcc atctatgtgt ataaggttct gaagcaggtc
     2821 caccctgaca ccggcatttc gtccaaggcc atgggcatca tgaactcgtt tgtgaacgac
     2881 attttcgagc gcatcgcagg tgaggcttcc cgcctggcgc attacaacaa gcgctcgacc
     2941 atcacctcca gggagatcca gacggccgtg cgcctgctgc tgcctgggga gttggccaag
     3001 cacgccgtgt ccgagggtac taaggccatc accaagtaca ccagcgctaa gtaaggcgcg
     3061 cccctgcagg gaattcgata tcataatcaa cctctggatt acaaaatttg tgaaagattg
     3121 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     3181 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     3241 ttagttcttg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     3301 cggctgttgg gcactgacaa ttccgtggct cgagagatct tcgactgtgc cttctagttg
     3361 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     3421 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     3481 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     3541 gcatgagatc tcacgtgcgg accgagcggc cgcaggaacc cctagtgatg gagttggcca
     3601 ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc
     3661 cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc
     3721 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca
     3781 accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     3841 cgtgaccgct acacttgcca gcgccttagc gcccgctcct ttcgctttct tcccttcctt
     3901 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     3961 ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg
     4021 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     4081 taatagtgga ctcttgttcc aaactggaac aacactcaac tctatctcgg gctattcttt
     4141 tgatttataa gggattttgc cgatttcggt ctattggtta aaaaatgagc tgatttaaca
     4201 aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag
     4261 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     4321 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     4381 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     4441 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc
     4501 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     4561 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     4621 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     4681 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     4741 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     4801 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     4861 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     4921 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     4981 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     5041 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     5101 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     5161 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     5221 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     5281 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     5341 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     5401 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     5461 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     5521 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     5581 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     5641 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     5701 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     5761 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     5821 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     5881 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     5941 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     6001 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     6061 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     6121 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     6181 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     6241 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     6301 tgctcacatg t
//
