LOCUS       sequence-212215-        5787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV expression of ChETA (hChR2 with E123T/H134R mutations) fused to 
            HA tag driven by hSynapsin promoter in Cre-ON manner for optogenetic
            activation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5787)
  AUTHORS   Li SJ, Vaughan A, Sturgill JF, Kepecs A
  TITLE     A Viral Receptor Complementation Strategy to Overcome CAV-2 Tropism 
            for Efficient Retrograde Targeting of Neurons.
  JOURNAL   Neuron. 2018 Jun 6;98(5):905-917.e5. doi: 
            10.1016/j.neuron.2018.05.028.
  PUBMED    29879392
REFERENCE   2  (bases 1 to 5787)
  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.2018.05"; journalName: "Neuron"; date: "2018-06-6-
            6"; volume: "98"; issue: "5"; pages: "905-917.e5"
COMMENT     Sequence Label: pAAV-hSyn-DIO {ChETA}on-WPRE
FEATURES             Location/Qualifiers
     source          1..5787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   2..131
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        176..623
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(661..694)
                     /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(745..778)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(791..817)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             complement(791..817)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(836..1762)
                     /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
                     PSMLYLATGHRVQWLRYATWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     regulatory      1759..1768
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1773..1806
                     /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    1857..1890
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    1915..2503
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1968..1988)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2386..2397)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2535..3011
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2734..2753)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3051..3191
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3051..3180
                     /label=AAV2 ITR
     rep_origin      3266..3721
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3353..3372)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3563..3584
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3738..3757)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3857..3879
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3917..3935)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4003..4107
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4108..4176,4177..4968)
                     /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(4326..4345)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5139..5727
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5628..5647
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcctgcaggc agctgcgcgc tcgctcgctc actgaggccg cccgggcgtc gggcgacctt
       61 tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca actccatcac
      121 taggggttcc tgcggccgca cgcgtgtgtc tagactgcag agggccctgc gtatgagtgc
      181 aagtgggttt taggaccagg atgaggcggg gtgggggtgc ctacctgacg accgaccccg
      241 acccactgga caagcaccca acccccattc cccaaattgc gcatccccta tcagagaggg
      301 ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc agcttcagca ccgcggacag
      361 tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct cagcactgaa ggcgcgctga
      421 cgtcactcgc cggtcccccg caaactcccc ttcccggcca ccttggtcgc gtccgcgccg
      481 ccgccggccc agccggaccg caccacgcga ggcgcgagat aggggggcac gggcgcgacc
      541 atctgcgctg cggcgccggc gactcagcgc tgcctcagtc tgcggtgggc agcggaggag
      601 tcgtgtcgtg cctgagagcg cagtcgagaa ggtaccggat cctctagagt cgactccgga
      661 ataacttcgt ataggatact ttatacgaag ttatgcagaa tggtagctgg attgtagctg
      721 ctattagcaa tatgaaacct cttaataact tcgtatagca tacattatac gaagttatgg
      781 cgcgtaatta ggcgtagtcg ggcacgtcgt aggggtacat ggtggcggcc gctggcacgg
      841 ctccggcctc ggcttcgtct tcgacgagag tctcgacctc gatctccgtt ccgccgatgt
      901 tcagtttggt ggttttgcgg atatctccgt gaatcaatat gtgctcgtgg atcaggacgc
      961 gcaggtagtg tcccaacaac ccccaacaat ttttactcat cagatcaata atcgtgtgac
     1021 ctacggtgga gccatagacg ctcaggacgc caaaaccttc gggccccaaa atgaagagaa
     1081 ttgggaacat accccagctc acgaaaaaca gccatgccat gccggtcacg acctggcggc
     1141 accgaccctt tggcacagta tgataaccct cgatatatgc tttggcggcg tgaaaaaatg
     1201 tgttcgcgcc atagcacaat ccaagacaaa agaagatgac tttaacatag ccggttgcca
     1261 tggcgctggt agccccccac acgatagtcc cgatgtctga gacaaggagt cccatggttc
     1321 tcctgctgta gtcgttgctc aggccggtga ggttgctcag gcggataagg atgacaggac
     1381 aagtgagcag ccaggttgca tagcgcagcc actgcacccg gtgtcctgtg gcaaggtaga
     1441 gcatagaggg attcttaaac tcaaaaaaga actcgagaat caccttaacc atttcaatgg
     1501 cgcacacata gatctcctcc cagccgcatg tagatttcca ggtttggtag gcatagaaca
     1561 tcagcagcaa aatgctgaat cctgctgcaa gccactgcag gacatttgac gcggtctgag
     1621 cgccgttcgt gccgcgagat tcaatccatc cggcacagta acattgatcc tcagggacca
     1681 ggacggaccc gttcaccacc acaggattag taacgaacaa aagttcgcgt ccgacggcag
     1741 acaaagcgcc gccatagtcc atggtggcta gcataacttc gtataaagta tcctatacga
     1801 agttatttgc cttaacccag aaattatcac tgttattctt tagaatggtg caaagaataa
     1861 cttcgtataa tgtatgctat acgaagttat gaattcgata tcaagcttat cgataatcaa
     1921 cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt
     1981 acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct
     2041 ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc
     2101 gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg
     2161 ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc
     2221 acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc
     2281 actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctat
     2341 gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca
     2401 gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt
     2461 cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcatcgata ccgagcgctg
     2521 ctcgagagat ctacgggtgg catccctgtg acccctcccc agtgcctctc ctggccctgg
     2581 aagttgccac tccagtgccc accagccttg tcctaataaa attaagttgc atcattttgt
     2641 ctgactaggt gtccttctat aatattatgg ggtggagggg ggtggtatgg agcaaggggc
     2701 aagttgggaa gacaacctgt agggcctgcg gggtctattg ggaaccaagc tggagtgcag
     2761 tggcacaatc ttggctcact gcaatctccg cctcctgggt tcaagcgatt ctcctgcctc
     2821 agcctcccga gttgttggga ttccaggcat gcatgaccag gctcagctaa tttttgtttt
     2881 tttggtagag acggggtttc accatattgg ccaggctggt ctccaactcc taatctcagg
     2941 tgatctaccc accttggcct cccaaattgc tgggattaca ggcgtgaacc actgctccct
     3001 tccctgtcct tctgattttg taggtaacca cgtgcggacc gagcggccgc aggaacccct
     3061 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
     3121 aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag
     3181 ctgcctgcag gggcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
     3241 ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt
     3301 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc cgctcctttc
     3361 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3421 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3481 ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     3541 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaactct
     3601 atctcgggct attcttttga tttataaggg attttgccga tttcggtcta ttggttaaaa
     3661 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt
     3721 ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
     3781 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     3841 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     3901 cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     3961 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     4021 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     4081 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     4141 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     4201 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     4261 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     4321 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     4381 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     4441 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     4501 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     4561 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     4621 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     4681 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     4741 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     4801 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     4861 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     4921 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     4981 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     5041 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     5101 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     5161 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     5221 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     5281 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     5341 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     5401 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     5461 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     5521 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     5581 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     5641 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     5701 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     5761 gccttttgct ggccttttgc tcacatg
//
