LOCUS       sequence-171064-        6384 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of ChR2-GFP for neuronal excitation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6384)
  AUTHORS   Boyden ES, Zhang F, Bamberg E, Nagel G, Deisseroth K
  TITLE     Millisecond-timescale, genetically targeted optical control of 
            neural activity.
  JOURNAL   Nat Neurosci. 2005 Sep . 8(9):1263-8.
  PUBMED    16116447
REFERENCE   2  (bases 1 to 6384)
  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: "Nat 
            Neurosci. 2005 Sep . 8(9):1263-8."
COMMENT     Sequence Label: AAV-CAG-ChR2-GFP
FEATURES             Location/Qualifiers
     source          1..6384
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   27..156
                     /label=AAV2 ITR
     promoter        467..744
                     /label=chicken beta-actin promoter
     primer_bind     1023..1045
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1107..1126
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1159..1168
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1165..2091
                     /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
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIHLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             1165..2091
                     /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
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIHLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             join(2113..2115,2116..2829)
                     /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             join(2113..2115,2116..2826)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2155..2176)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2416..2435)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2763..2784
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2854..3442
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2907..2927)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3325..3336)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3485..3606
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3496..3515)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3550..3569
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3768..3897
                     /label=AAV2 ITR
     rep_origin      complement(4134..4722)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4214..4233)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4893..5684,5685..5753))
                     /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     5516..5535
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5754..5858)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5884..6339)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6021..6042)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6233..6252
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 tgcgggcctc ttcgctatta cgccagctgc gcgctcgctc gctcactgag gccgcccggg
       61 caaagcccgg gcgtcgggcg acctttggtc gcccggcctc agtgagcgag cgagcgcgca
      121 gagagggagt ggccaactcc atcactaggg gttccttgta gttaatgatt aacccgccat
      181 gctacttatc tacgtagcca tgctctagga agagtaccat tgacgtcaat aatgacgtat
      241 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
      301 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
      361 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
      421 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtcga ggtgagcccc
      481 acgttctgct tcactctccc catctccccc ccctccccac ccccaatttt gtatttattt
      541 attttttaat tattttgtgc agcgatgggg gcgggggggg ggggggggcg cgcgccgggc
      601 ggggcggggc ggggcgaggg gcggggcggg gcgaggcgga gaggtgcggc ggcagccaat
      661 cagagcggcg cgctccgaaa gtttcctttt atggcgaggc ggcggcggcg gcggccctat
      721 aaaaagcgaa gcgcgcggcg ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc
      781 tccgccgccg cctcgcgccg cccgccccgg ctctgactga ccgcgttact cccacaggtg
      841 agcgggcggg acggcccttc tcctccgggc tgtaattagc gcttggttta atgacggctt
      901 gtttcttttc tgtggctgcg tgaaagcctt gaggggctcc gggagggccc tttgtgcggg
      961 gggagcggct cggggctgtc cgcgggggga cggctgcctt cgggggggac ggggcagggc
     1021 ggggttcggc ttctggcgtg tgaccggcgg ctctagagcc tctgctaacc atgttcatgc
     1081 cttcttcttt ttcctacagc tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt
     1141 tggcaaagaa ttggatccgc caccatggac tatggcggcg ctttgtctgc cgtcggacgc
     1201 gaacttttgt tcgttactaa tcctgtggtg gtgaacgggt ccgtcctggt ccctgaggat
     1261 caatgttact gtgccggatg gattgaatct cgcggcacga acggcgctca gaccgcgtca
     1321 aatgtcctgc agtggcttgc agcaggattc agcattttgc tgctgatgtt ctatgcctac
     1381 caaacctgga aatctacatg cggctgggag gagatctatg tgtgcgccat tgaaatggtt
     1441 aaggtgattc tcgagttctt ttttgagttt aagaatccct ctatgctcta ccttgccaca
     1501 ggacaccggg tgcagtggct gcgctatgca gagtggctgc tcacttgtcc tgtcatcctt
     1561 atccacctga gcaacctcac cggcctgagc aacgactaca gcaggagaac catgggactc
     1621 cttgtctcag acatcgggac tatcgtgtgg ggggctacca gcgccatggc aaccggctat
     1681 gttaaagtca tcttcttttg tcttggattg tgctatggcg cgaacacatt ttttcacgcc
     1741 gccaaagcat atatcgaggg ttatcatact gtgccaaagg gtcggtgccg ccaggtcgtg
     1801 accggcatgg catggctgtt tttcgtgagc tggggtatgt tcccaattct cttcattttg
     1861 gggcccgaag gttttggcgt cctgagcgtc tatggctcca ccgtaggtca cacgattatt
     1921 gatctgatga gtaaaaattg ttgggggttg ttgggacact acctgcgcgt cctgatccac
     1981 gagcacatat tgattcacgg agatatccgc aaaaccacca aactgaacat cggcggaacg
     2041 gagatcgagg tcgagactct cgtcgaagac gaagccgagg ccggagccgt gccagcggca
     2101 ccggtagtag cagtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc
     2161 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat
     2221 gccacctacg gcaagctgac cctgaagttc atttgcacca ccggcaagct gcccgtgccc
     2281 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac
     2341 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
     2401 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc
     2461 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc
     2521 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag
     2581 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg
     2641 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
     2701 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat
     2761 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg
     2821 tacaagtaag aattcgatat caagcttatc gataatcaac ctctggatta caaaatttgt
     2881 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     2941 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     3001 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     3061 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     3121 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     3181 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     3241 tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc
     3301 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     3361 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     3421 tccctttggg ccgcctcccc gcatcgatac cgtcgacccg ggcggccgct tcgagcagac
     3481 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
     3541 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     3601 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag
     3661 gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tcttcctaga
     3721 gcatggctac gtagataagt agcatggcgg gttaatcatt aactacaagg aacccctagt
     3781 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     3841 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     3901 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     3961 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     4021 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     4081 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4141 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4201 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4261 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4321 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4381 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4441 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4501 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4561 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4621 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4681 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4741 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4801 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4861 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4921 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4981 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     5041 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     5101 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     5161 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     5221 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5281 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5341 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5401 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5461 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5521 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5581 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5641 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5701 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5761 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5821 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     5881 cctaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     5941 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     6001 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     6061 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     6121 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     6181 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     6241 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     6301 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca ttcaggctgc
     6361 gcaactgttg ggaagggcga tcgg
//
