LOCUS       Exported                7022 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV expression of humanized ChR2(H134R) fused to mCherry driven by 
            CaMKIIa promoter for optogenetic activation.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-CaMKIIa-hChR2(H134R)-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7022)
  TITLE     hChR2 plasmids
REFERENCE   2  (bases 1 to 7022)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7022
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     64..83
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   245..385
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   256..385
                     /label=AAV2 ITR
     polyA_signal    425..901
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     683..702
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     misc_feature    933..1521
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1039..1050
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1448..1468
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1546..2253)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1824..1843)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2093..2111
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2266..3192)
                     /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"
     regulatory      3189..3198
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     promoter        complement(3233..4521)
                     /label=CaMKII promoter
                     /note="promoter for the mouse calcium/calmodulin-dependent 
                     kinase II alpha subunit"
     repeat_region   4540..4669
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(4731..5319)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4811..4830)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5490..6350)
                     /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     6113..6132
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6351..6455)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6523..6541
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6579..6601)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6701..6720
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(6737..170)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6874..6895)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa
       61 ggaagggaag aaagcgaaag gagcgggcgc taaggcgctg gcaagtgtag cggtcacgct
      121 gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt actatggttg
      181 ctttgacgta tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
      241 cgcccctgca ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg
      301 cgtcgggcga cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg
      361 gccaactcca tcactagggg ttcctgcggc cgctcggtcc gcacgtggtt acctacaaaa
      421 tcagaaggac agggaaggga gcagtggttc acgcctgtaa tcccagcaat ttgggaggcc
      481 aaggtgggta gatcacctga gattaggagt tggagaccag cctggccaat atggtgaaac
      541 cccgtctcta ccaaaaaaac aaaaattagc tgagcctggt catgcatgcc tggaatccca
      601 acaactcggg aggctgaggc aggagaatcg cttgaaccca ggaggcggag attgcagtga
      661 gccaagattg tgccactgca ctccagcttg gttcccaata gaccccgcag gccctacagg
      721 ttgtcttccc aacttgcccc ttgctccata ccacccccct ccaccccata atattataga
      781 aggacaccta gtcagacaaa atgatgcaac ttaattttat taggacaagg ctggtgggca
      841 ctggagtggc aacttccagg gccaggagag gcactgggga ggggtcacag ggatgccacc
      901 cgtagatctc tcgagcagcg ctcggtatcg atgcggggag gcggcccaaa gggagatccg
      961 actcgtctga gggcgaaggc gaagacgcgg aagaggccgc agagccggca gcaggccgcg
     1021 ggaaggaagg tccgctggat tgagggccga agggacgtag cagaaggacg tcccgcgcag
     1081 aatccaggtg gcaacatagg cgagcagcca aggaaaggac gatgatttcc ccgacaacac
     1141 cacggaattg tcagtgccca acagccgagc ccctgtccag cagcgggcaa ggcaggcggc
     1201 gatgagttcc gccgtggcaa tagggagggg gaaagcgaaa gttccggaaa ggagctgaca
     1261 ggtggtggca atgccccaac cagtgggggt tgcgtcagca aacacagtgc acaccacgcc
     1321 acgttgcctg acaacgggcc acaactcctc ataaagagac agcaaccagg atttatacaa
     1381 ggaggagaaa atgaaagcca tacgggaagc aatagcatga tacaaaggca ttaaagcagc
     1441 gtatccacat agcgtaaaag gagcaacata gttaagaata ccagtcaatc tttcacaaat
     1501 tttgtaatcc agaggttgat tatcgataag cttgatatcg aattcttact tgtacagctc
     1561 gtccatgccg ccggtggagt ggcggccctc ggcgcgttcg tactgttcca cgatggtgta
     1621 gtcctcgttg tgggaggtga tgtccaactt gatgttgacg ttgtaggcgc cgggcagctg
     1681 cacgggcttc ttggccttgt aggtggtctt gacctcagcg tcgtagtggc cgccgtcctt
     1741 cagcttcagc ctctgcttga tctcgccctt cagggcgccg tcctcggggt acatccgctc
     1801 ggaggaggcc tcccagccca tggtcttctt ctgcattacg gggccgtcgg aggggaagtt
     1861 ggtgccgcgc agcttcacct tgtagatgaa ctcgccgtcc tgcagggagg agtcctgggt
     1921 cacggtcacc acgccgccgt cctcgaagtt catcacgcgc tcccacttga agccctcggg
     1981 gaaggacagc ttcaagtagt cggggatgtc ggcggggtgc ttcacgtagg ccttggagcc
     2041 gtacatgaac tgaggggaca ggatgtccca ggcgaagggc agggggccac ccttggtcac
     2101 cttcagcttg gcggtctggg tgccctcgta ggggcggccc tcgccctcgc cctcgatctc
     2161 gaactcgtgg ccgttcacgg agccctccat gtgcaccttg aagcgcatga actccttgat
     2221 gatggccatg ttatcctcct cgcccttgct cacggcggcc gctggcacgg ctccggcctc
     2281 ggcttcgtct tcgacgagag tctcgacctc gatctccgtt ccgccgatgt tcagtttggt
     2341 ggttttgcgg atatctccgt gaatcaatat gtgctcgtgg atcaggacgc gcaggtagtg
     2401 tcccaacaac ccccaacaat ttttactcat cagatcaata atcgtgtgac ctacggtgga
     2461 gccatagacg ctcaggacgc caaaaccttc gggccccaaa atgaagagaa ttgggaacat
     2521 accccagctc acgaaaaaca gccatgccat gccggtcacg acctggcggc accgaccctt
     2581 tggcacagta tgataaccct cgatatatgc tttggcggcg tgaaaaaatg tgttcgcgcc
     2641 atagcacaat ccaagacaaa agaagatgac tttaacatag ccggttgcca tggcgctggt
     2701 agccccccac acgatagtcc cgatgtctga gacaaggagt cccatggttc tcctgctgta
     2761 gtcgttgctc aggccggtga ggttgctcag gcggataagg atgacaggac aagtgagcag
     2821 ccactctgca tagcgcagcc actgcacccg gtgtcctgtg gcaaggtaga gcatagaggg
     2881 attcttaaac tcaaaaaaga actcgagaat caccttaacc atttcaatgg cgcacacata
     2941 gatctcctcc cagccgcatg tagatttcca ggtttggtag gcatagaaca tcagcagcaa
     3001 aatgctgaat cctgctgcaa gccactgcag gacatttgac gcggtctgag cgccgttcgt
     3061 gccgcgagat tcaatccatc cggcacagta acattgatcc tcagggacca ggacggaccc
     3121 gttcaccacc acaggattag taacgaacaa aagttcgcgt ccgacggcag acaaagcgcc
     3181 gccatagtcc atggtggcct accggtaccc ggggatccgg taccgctcta gagctgcccc
     3241 cagaactagg ggccactcgc ctgcccgtgc tcctgagtgc aaacggagaa ccggcttgac
     3301 tgacgagctt ggggcttctg agcagggcac tgtggctgct cacagcctcg cgagccttcg
     3361 tcagcatcca ggtccccatg gcaaccacct ccgaaacgcc ctgttcccgt tgccccggta
     3421 actgcctccc caacacctgc ctgcctttca ctttaaaacc tgctcctctt tgccctggcc
     3481 ttatatacac aatactgcag gactgtgtgg gcccaggagc aagtggaccc tgttccccca
     3541 gggtgatgta ctgagggggt tggagaaggg gatgcgaaca aacttagtcc acaagtgatc
     3601 atcgctcttc ctacctcccc cacctccaac tatagggccc ttttaacaga cgggatggga
     3661 ttaagtgggg caaagaggca gttgctatgg taacggctag tgggtgccac attctgggat
     3721 ggtccttgag aaagacagga tgtagttacc ctggcaactt catctcctct gaagagacag
     3781 gtattgcccc gttgccctgg caacagctgg gttctaaggt ttacagggga agctaattca
     3841 tctctccttc ccttccccct tgtagtctct gtaatggttt ctttgcttcc tctgggaatt
     3901 ggtctggttg acttggcctc tgtctctgtc cttgtttctt tccagctctc cctgcttttt
     3961 agccattgtg catatcacac cgccacctta gcgtgagaag aagtaccaaa cagacccaga
     4021 atgctgacgg aatgactctg tctgctttct cacccctatc tgtcaccact gagcagcagg
     4081 gcccctcctg aggcctctgc cacagctctc cccacccatc ctcccagagg gaccatgaag
     4141 atttggtctc ctggggctcc tgtcccagtt tctgcttgtg cttcccgtca aggtaggact
     4201 ggagagaggc aactataatg aggtgaacgt ggctggactc acccagggct gggcgggtgg
     4261 tgtagatgta ccctgccctg gactcttctt tcctaaactc attggtcatc tgccctaatg
     4321 agttcactcc ttctgtatat cgtacttgca tgcccccaag gcatggaatt taagattagt
     4381 aatctgagtt caagttctag ctctgacatt tctgatgtga ccctgggcag gtcactgagg
     4441 aagctctctg cacccccatc tcattttcta gaaaatcaga agaagaaccc catggagatg
     4501 aagtgaccta aggccataat gttaaacgcg tgcggccgca ggaaccccta gtgatggagt
     4561 tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc
     4621 gacgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcagga catgtgagca
     4681 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4741 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4801 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4861 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4921 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4981 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5041 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5101 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5161 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5221 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5281 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5341 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5401 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5461 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5521 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5581 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5641 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5701 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5761 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5821 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5881 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5941 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6001 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6061 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6121 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6181 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6241 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6301 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6361 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6421 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6481 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     6541 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     6601 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     6661 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
     6721 ctgagagtgc accataaaat tgtaaacgtt aatattttgt taaaattcgc gttaaatttt
     6781 tgttaaatca gctcattttt taaccaatag accgaaatcg gcaaaatccc ttataaatca
     6841 aaagaatagc ccgagataga gttgagtgtt gttccagttt ggaacaagag tccactatta
     6901 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     6961 cgtgaaccat cacccaaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     7021 aa
//
