LOCUS       sequence-81429-e        7012 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Historical plasmid. For inhibition of neuronal activity, note that 
            Addgene Plasmid #66709 is a light-gated chloride channel with 
            improved anion selectivity (iChloC).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7012)
  AUTHORS   Wietek J, Wiegert JS, Adeishvili N, Schneider F, Watanabe H, Tsunoda
            SP, Vogt A, Elstner M, Oertner TG, Hegemann P
  TITLE     Conversion of Channelrhodopsin into a Light-Gated Chloride Channel.
  JOURNAL   Science. 2014 Mar 27.
  PUBMED    24674867
REFERENCE   2  (bases 1 to 7012)
  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: "Science. 
            2014 Mar 27."
COMMENT     Sequence Label: pAAV Syn ChR E90R-D156N-T159C 2A tDimer
FEATURES             Location/Qualifiers
     source          1..7012
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        27..474
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      494..503
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             500..1426
                     /codon_start=1
                     /product="N-terminal fragment of channelrhodopsin-2 from 
                     the alga Chlamydomonas reinhardtii (Nagel et al., 2003)"
                     /label=ChR2
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIRMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIHLSNLTGLSNDYSRRTMGLLVSNIGCIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             1436..1489
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     regulatory      1505..1514
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1509..1518
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1511..2188
                     /codon_start=1
                     /product="rapidly maturing tetrameric variant of DsRed 
                     fluorescent protein (Bevis and Glick, 2002)"
                     /label=DsRed-Express
                     /note="mammalian codon-optimized"
                     /translation="MVASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     VVTVTQDSSLQDGTLIYKVKFRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFL"
     CDS             2231..2905
                     /codon_start=1
                     /product="rapidly maturing tetrameric variant of DsRed 
                     fluorescent protein (Bevis and Glick, 2002)"
                     /label=DsRed-Express
                     /note="mammalian codon-optimized"
                     /translation="ASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAKL
                     KVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGTLIYKVKFRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIHQA
                     LKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRHHL
                     FL"
     misc_feature    2933..3521
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2986..3006)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3404..3415)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3524..3540)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3525..3541)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    3601..4077
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3800..3819)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4117..4257
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4117..4246
                     /label=AAV2 ITR
     rep_origin      4332..4787
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4419..4438)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4629..4650
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4804..4823)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4923..4945
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4983..5001)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5069..5173
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5174..5242,5243..6034)
                     /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(5392..5411)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6205..6793
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6694..6713
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   6855..6995
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6866..6995
                     /label=AAV2 ITR
ORIGIN
        1 ctagactgca gagggccctg cgtatgagtg caagtgggtt ttaggaccag gatgaggcgg
       61 ggtgggggtg cctacctgac gaccgacccc gacccactgg acaagcaccc aacccccatt
      121 ccccaaattg cgcatcccct atcagagagg gggaggggaa acaggatgcg gcgaggcgcg
      181 tgcgcactgc cagcttcagc accgcggaca gtgccttcgc ccccgcctgg cggcgcgcgc
      241 caccgccgcc tcagcactga aggcgcgctg acgtcactcg ccggtccccc gcaaactccc
      301 cttcccggcc accttggtcg cgtccgcgcc gccgccggcc cagccggacc gcaccacgcg
      361 aggcgcgaga taggggggca cgggcgcgac catctgcgct gcggcgccgg cgactcagcg
      421 ctgcctcagt ctgcggtggg cagcggagga gtcgtgtcgt gcctgagagc gcagtcgaga
      481 attcaagctg ctagccacca tggattatgg aggcgccctg agtgccgttg ggcgcgagct
      541 gctatttgta acgaacccag tagtcgtcaa tggctctgta cttgtgcctg aggaccagtg
      601 ttactgcgcg ggctggattg agtcgcgtgg cacaaacggt gcccaaacgg cgtcgaacgt
      661 gctgcaatgg cttgctgctg gcttctccat cctactgctt atgttttacg cctaccaaac
      721 atggaagtca acctgcggct gggaggagat ctatgtgtgc gctatcagga tggtcaaggt
      781 gattctcgag ttcttcttcg agtttaagaa cccgtccatg ctgtatctag ccacaggcca
      841 ccgcgtccag tggttgcgtt acgccgagtg gcttctcacc tgcccggtca ttctcattca
      901 cctgtcaaac ctgacgggct tgtccaacga ctacagcagg cgcaccatgg gtctgcttgt
      961 gtctaatatt ggctgtattg tgtggggcgc cacttccgcc atggccaccg gatacgtcaa
     1021 ggtcatcttc ttctgcctgg gtctgtgtta tggtgctaac acgttctttc acgctgccaa
     1081 ggcctacatc gagggttacc acaccgtgcc gaagggccgg tgtcgccagg tggtgactgg
     1141 catggcttgg ctcttcttcg tatcatgggg tatgttcccc atcctgttca tcctcggccc
     1201 cgagggcttc ggcgtcctga gcgtgtacgg ctccaccgtc ggccacacca tcattgacct
     1261 gatgtcgaag aactgctggg gtctgctcgg ccactacctg cgcgtgctga tccacgagca
     1321 tatcctcatc cacggcgaca ttcgcaagac caccaaattg aacattggtg gcactgagat
     1381 tgaggtcgag acgctggtgg aggacgaggc cgaggctggc gcggtacccg cggccgaggg
     1441 cagaggaagt cttctaacat gcggtgacgt ggaggagaat cccggccctg caccgggatc
     1501 cgcggccgcc atggtggcct cctccgagga cgtcatcaaa gagttcatgc gcttcaaggt
     1561 gcgcatggag ggctccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg
     1621 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggcggcc ccctgccctt
     1681 cgcctgggac atcctgtccc cccagttcca gtacggctcc aaggcgtacg tgaagcaccc
     1741 cgccgacatc cccgactaca agaagctgtc cttccccgag ggcttcaagt gggagcgcgt
     1801 gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg
     1861 cacgctgatc tacaaggtga agttccgcgg caccaacttc ccccccgacg gccccgtaat
     1921 gcagaagaag accatgggct gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt
     1981 gctgaagggc gagatccacc aggccctgaa gctgaaggac ggcggccact acctggtgga
     2041 gttcaagacc atctacatgg ccaagaagcc cgtgcagctg cccggctact actacgtgga
     2101 caccaagctg gacatcacct cccacaacga ggactacacc atcgtggaac agtacgagcg
     2161 ctccgagggc cgccaccacc tgttcctggg gcatggcacc ggcagcaccg gcagcggcag
     2221 ctccggcacc gcctcctccg aggacgtcat caaagagttc atgcgcttca aggtgcgcat
     2281 ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta
     2341 cgagggcacc cagaccgcca agctgaaggt gaccaagggc ggccccctgc ccttcgcctg
     2401 ggacatcctg tccccccagt tccagtacgg ctccaaggcg tacgtgaagc accccgccga
     2461 catccccgac tacaagaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa
     2521 cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcacgct
     2581 gatctacaag gtgaagttcc gcggcaccaa cttccccccc gacggccccg taatgcagaa
     2641 gaagaccatg ggctgggagg cctccaccga gcgcctgtac ccccgcgacg gcgtgctgaa
     2701 gggcgagatc caccaggccc tgaagctgaa ggacggcggc cactacctgg tggagttcaa
     2761 gaccatctac atggccaaga agcccgtgca gctgcccggc tactactacg tggacaccaa
     2821 gctggacatc acctcccaca acgaggacta caccatcgtg gaacagtacg agcgctccga
     2881 gggccgccac cacctgttcc tgtaggtcga cttcgagcaa aagcttatcg ataatcaacc
     2941 tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac
     3001 gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt
     3061 cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt
     3121 tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg
     3181 cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac
     3241 ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac
     3301 tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt
     3361 tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc
     3421 ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg
     3481 ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc gtcgacctcg
     3541 agacgcgatc gcggcgcgcc ggcctctctg gcctcgacct cgagtctaga gatgatctac
     3601 gggtggcatc cctgtgaccc ctccccagtg cctctcctgg ccctggaagt tgccactcca
     3661 gtgcccacca gccttgtcct aataaaatta agttgcatca ttttgtctga ctaggtgtcc
     3721 ttctataata ttatggggtg gaggggggtg gtatggagca aggggcaagt tgggaagaca
     3781 acctgtaggg cctgcggggt ctattgggaa ccaagctgga gtgcagtggc acaatcttgg
     3841 ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc tgcctcagcc tcccgagttg
     3901 ttgggattcc aggcatgcat gaccaggctc agctaatttt tgtttttttg gtagagacgg
     3961 ggtttcacca tattggccag gctggtctcc aactcctaat ctcaggtgat ctacccacct
     4021 tggcctccca aattgctggg attacaggcg tgaaccactg ctcccttccc tgtccttctg
     4081 attttgtagg taaccacgtg cggaccgagc ggccgcagga acccctagtg atggagttgg
     4141 ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac
     4201 gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc
     4261 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa
     4321 gcaaccatag tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     4381 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     4441 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     4501 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc
     4561 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     4621 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cgggctattc
     4681 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     4741 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct
     4801 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc
     4861 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     4921 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa
     4981 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac
     5041 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     5101 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     5161 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5221 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5281 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     5341 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     5401 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     5461 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     5521 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     5581 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5641 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5701 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     5761 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     5821 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     5881 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     5941 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     6001 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     6061 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     6121 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     6181 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6241 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6301 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt
     6361 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     6421 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     6481 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     6541 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     6601 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     6661 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     6721 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     6781 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     6841 ttttgctcac atgtcctgca ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc
     6901 aaagcccggg cgtcgggcga cctttggtcg cccggcctca gtgagcgagc gagcgcgcag
     6961 agagggagtg gccaactcca tcactagggg ttcctgcggc cgcacgcgtc ga
//
