LOCUS       sequence-399615-        6602 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An adeno-associated viral vector expressing channelrhodopsin-2 fused
            to a V5 epitope and the ligand-binding domain of estrogen receptor 
            (ChRERa) from a synapsin promoter, for use in PET imaging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6602)
  AUTHORS   Bonaventura J, Boehm MA, Jedema HP, Solis O, Pignatelli M, Song X, 
            Lu H, Richie CT, Zhang S, Gomez JL, Lam S, Morales M, Gharbawie OA, 
            Pomper MG, Stein EA, Bradberry CW, Michaelides M
  TITLE     Expression of the excitatory opsin ChRERalpha can be traced 
            longitudinally in rat and nonhuman primate brains with PET imaging.
  JOURNAL   Sci Transl Med. 2023 Jul 26;15(706):eadd1014. doi: 
            10.1126/scitranslmed.add1014. Epub 2023 Jul 26.
  PUBMED    37494470
REFERENCE   2  (bases 1 to 6602)
  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"; journalName: "Sci Transl 
            Med."; date: "2023-07-26"; volume: "15(706):eadd1014. doi"; pages: "
            10.1126/scitranslmed.add1014. Epub 2023 Jul 26"
COMMENT     Sequence Label: pOTTC1784 - pAAV SYN1 hChR2(H134R)-V5-ER-LBD
            (...
FEATURES             Location/Qualifiers
     source          1..6602
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        175..622
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             660..1586
                     /codon_start=1
                     /product="H134R mutant of an N-terminal fragment of 
                     channelrhodopsin-2 from the alga Chlamydomonas reinhardtii 
                     (Nagel et al., 2005)"
                     /label=ChR2(H134R)
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             1653..1694
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             1740..2675
                     /codon_start=1
                     /product="mutated ligand-binding domain of the human 
                     estrogen receptor (Feil et al., 1997)"
                     /label=ERT2
                     /translation="AGDMRAANLWPSPLMIKRSKKNSLALSLTADQMVSALLDAEPPIL
                     YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEI
                     LMIGLVWRSMEHPGKLLFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEF
                     VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL
                     AQLLLILSHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEMLDAHRLHAPTSRGGASVEET
                     DQSHLATAGSTSSHSLQKYYITGEAEGFPAT"
     CDS             1788..2681
                     /codon_start=1
                     /gene="ESR1"
                     /product="estrogen nuclear receptor alpha ligand-binding 
                     domain"
                     /label=ER-LBD
                     /translation="KRSKKNSLALSLTADQMVSALLDAEPPILYSEYDPTRPFSEASMM
                     GLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEILMIGLVWRSMEHPGKL
                     LFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEFVCLKSIILLNSGVYTF
                     LSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRLAQLLLILSHIRHMSNK
                     GMEHLYSMKCKNVVPLYDLLLEMLDAHRLHAPTSRGGASVEETDQSHLATAGSTSSHSL
                     QKYYITGEAEGFPATV"
     misc_feature    2729..3317
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2782..2802)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3200..3211)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3349..3825
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3548..3567)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3865..4005
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3865..3994
                     /label=AAV2 ITR
     rep_origin      4080..4535
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4167..4186)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4377..4398
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4552..4571)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4671..4693
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4731..4749)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4817..4921
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4922..4990,4991..5782)
                     /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(5140..5159)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5953..6541
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6442..6461
                     /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 gcgtgtgtct agactgcaga gggccctgcg tatgagtgca
      181 agtgggtttt aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga
      241 cccactggac aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg
      301 gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt
      361 gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac
      421 gtcactcgcc ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc
      481 cgccggccca gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca
      541 tctgcgctgc ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt
      601 cgtgtcgtgc ctgagagcgc agtcgagaag gtaccggatc cgaagttatc gctagcacca
      661 tggactatgg cggcgctttg tctgccgtcg gacgcgaact tttgttcgtt actaatcctg
      721 tggtggtgaa cgggtccgtc ctggtccctg aggatcaatg ttactgtgcc ggatggattg
      781 aatctcgcgg cacgaacggc gctcagaccg cgtcaaatgt cctgcagtgg cttgcagcag
      841 gattcagcat tttgctgctg atgttctatg cctaccaaac ctggaaatct acatgcggct
      901 gggaggagat ctatgtgtgc gccattgaaa tggttaaggt gattctcgag ttcttttttg
      961 agtttaagaa tccctctatg ctctaccttg ccacaggaca ccgggtgcag tggctgcgct
     1021 atgcagagtg gctgctcact tgtcctgtca tccttatccg cctgagcaac ctcaccggcc
     1081 tgagcaacga ctacagcagg agaaccatgg gactccttgt ctcagacatc gggactatcg
     1141 tgtggggggc taccagcgcc atggcaaccg gctatgttaa agtcatcttc ttttgtcttg
     1201 gattgtgcta tggcgcgaac acattttttc acgccgccaa agcatatatc gagggttatc
     1261 atactgtgcc aaagggtcgg tgccgccagg tcgtgaccgg catggcatgg ctgtttttcg
     1321 tgagctgggg tatgttccca attctcttca ttttggggcc cgaaggtttt ggcgtcctga
     1381 gcgtctatgg ctccaccgta ggtcacacga ttattgatct gatgagtaaa aattgttggg
     1441 ggttgttggg acactacctg cgcgtcctga tccacgagca catattgatt cacggagata
     1501 tccgcaaaac caccaaactg aacatcggcg gaacggagat cgaggtcgag actctcgtcg
     1561 aagacgaagc cgaggccgga gccgtgccag cggccgccac cggtggatct atggaagctt
     1621 actgtcatat taaggggttc gatgccgtga atggaaagcc aatacccaat cctctgctgg
     1681 gcttggactc aactagaacc ggcggcggtt gtccatgggc cactggttca ggttcatctg
     1741 ctggagacat gagagctgcc aacctttggc caagcccgct catgatcaaa cgctctaaga
     1801 agaacagcct ggccttgtcc ctgacggccg accagatggt cagtgccttg ttggatgctg
     1861 agccccccat actctattcc gagtatgatc ctaccagacc cttcagtgaa gcctccatga
     1921 tgggcttact gaccaacctg gcagacaggg agctggttca catgatcaac tgggcgaaga
     1981 gggtgccagg ctttgtggat ttgaccctcc atgatcaggt ccaccttctg gaatgtgcct
     2041 ggctagagat cctgatgatt ggcctcgtct ggcgctccat ggagcaccca gggaagctac
     2101 tgtttgctcc taacttgctc ttggacagga accagggaaa atgtgtagag ggcatggtgg
     2161 agatcttcga catgctgctg gctacatcat ctcggttccg catgatgaat ctgcagggag
     2221 aggagtttgt gtgcctcaaa tctattattt tgcttaattc tggagtgtac acatttctgt
     2281 ccagcaccct gaagtctctg gaagagaagg accatatcca ccgagtcctg gacaagatca
     2341 cagacacttt gatccacctg atggccaagg caggcctgac cctgcagcag cagcaccagc
     2401 ggctggccca gctcctcctc atcctctccc acatcaggca catgagtaac aaaggcatgg
     2461 agcatctgta cagcatgaag tgcaagaacg tggtgcccct ctatgacctg ctgctggaga
     2521 tgctggacgc ccaccgccta catgcgccca ctagccgtgg aggggcatcc gtggaggaga
     2581 cggaccaaag ccacttggcc actgcgggct ctacttcatc gcattccttg caaaagtatt
     2641 acatcacggg ggaggcagag ggtttccctg ccacagtcta ggcaggtggg cgcgccataa
     2701 cttcgaattc gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag
     2761 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     2821 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     2881 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     2941 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     3001 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     3061 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     3121 gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc acctggattc tgcgcgggac
     3181 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     3241 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     3301 ttgggccgcc tccccgcatc gataccgagc gctgctcgag agatctacgg gtggcatccc
     3361 tgtgacccct ccccagtgcc tctcctggcc ctggaagttg ccactccagt gcccaccagc
     3421 cttgtcctaa taaaattaag ttgcatcatt ttgtctgact aggtgtcctt ctataatatt
     3481 atggggtgga ggggggtggt atggagcaag gggcaagttg ggaagacaac ctgtagggcc
     3541 tgcggggtct attgggaacc aagctggagt gcagtggcac aatcttggct cactgcaatc
     3601 tccgcctcct gggttcaagc gattctcctg cctcagcctc ccgagttgtt gggattccag
     3661 gcatgcatga ccaggctcag ctaatttttg tttttttggt agagacgggg tttcaccata
     3721 ttggccaggc tggtctccaa ctcctaatct caggtgatct acccaccttg gcctcccaaa
     3781 ttgctgggat tacaggcgtg aaccactgct cccttccctg tccttctgat tttgtaggta
     3841 accacgtgcg gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc
     3901 tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg
     3961 cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt
     4021 attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta
     4081 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     4141 tacacttgcc agcgccttag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     4201 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     4261 tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc
     4321 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     4381 actcttgttc caaactggaa caacactcaa ctctatctcg ggctattctt ttgatttata
     4441 agggattttg ccgatttcgg tctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     4501 cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg
     4561 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     4621 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     4681 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
     4741 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     4801 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     4861 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     4921 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     4981 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     5041 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     5101 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     5161 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     5221 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     5281 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     5341 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     5401 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     5461 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     5521 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     5581 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     5641 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     5701 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     5761 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     5821 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     5881 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     5941 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     6001 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     6061 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     6121 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     6181 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     6241 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     6301 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     6361 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     6421 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     6481 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     6541 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     6601 gt
//
