LOCUS       Exported                7031 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV expression of CaMKII-driven stabilized step function opsin 
            (SSFO) for optogenetic activation..
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-CaMKIIa-hChR2(C128S/D156A)-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7031)
  AUTHORS   Yizhar O, Fenno LE, Prigge M, Schneider F, Davidson TJ, O'Shea DJ, 
            Sohal VS, Goshen I, Finkelstein J, Paz JT, Stehfest K, Fudim R, 
            Ramakrishnan C, Huguenard JR, Hegemann P, Deisseroth K
  TITLE     Neocortical excitation/inhibition balance in information processing 
            and social dysfunction.
  JOURNAL   Nature. 2011 Jul 27;477(7363):171-8. doi: 10.1038/nature10360.
  PUBMED    21796121
REFERENCE   2  (bases 1 to 7031)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7031
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        160..1448
                     /label=CaMKII promoter
                     /note="promoter for the mouse calcium/calmodulin-dependent 
                     kinase II alpha subunit"
     regulatory      1481..1490
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1487..2413
                     /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
                     PSMLYLATGHRVQWLRYAEWLLTSPVILIHLSNLTGLSNDYSRRTMGLLVSAIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             2426..3133
                     /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(2568..2586)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2836..2855
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    3158..3746
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3211..3231)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3629..3640)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3778..4254
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3977..3996)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4294..4434
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4294..4423
                     /label=AAV2 ITR
     rep_origin      4509..4964
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4596..4615)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4806..4827
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4981..5000)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5100..5122
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5160..5178)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5246..5350
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5351..6211
                     /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(5569..5588)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6382..6970
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6871..6890
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtttaac attatggcct taggtcactt
      181 catctccatg gggttcttct tctgattttc tagaaaatga gatgggggtg cagagagctt
      241 cctcagtgac ctgcccaggg tcacatcaga aatgtcagag ctagaacttg aactcagatt
      301 actaatctta aattccatgc cttgggggca tgcaagtacg atatacagaa ggagtgaact
      361 cattagggca gatgaccaat gagtttagga aagaagagtc cagggcaggg tacatctaca
      421 ccacccgccc agccctgggt gagtccagcc acgttcacct cattatagtt gcctctctcc
      481 agtcctacct tgacgggaag cacaagcaga aactgggaca ggagccccag gagaccaaat
      541 cttcatggtc cctctgggag gatgggtggg gagagctgtg gcagaggcct caggaggggc
      601 cctgctgctc agtggtgaca gataggggtg agaaagcaga cagagtcatt ccgtcagcat
      661 tctgggtctg tttggtactt cttctcacgc taaggtggcg gtgtgatatg cacaatggct
      721 aaaaagcagg gagagctgga aagaaacaag gacagagaca gaggccaagt caaccagacc
      781 aattcccaga ggaagcaaag aaaccattac agagactaca agggggaagg gaaggagaga
      841 tgaattagct tcccctgtaa accttagaac ccagctgttg ccagggcaac ggggcaatac
      901 ctgtctcttc agaggagatg aagttgccag ggtaactaca tcctgtcttt ctcaaggacc
      961 atcccagaat gtggcaccca ctagccgtta ccatagcaac tgcctctttg ccccacttaa
     1021 tcccatcccg tctgttaaaa gggccctata gttggaggtg ggggaggtag gaagagcgat
     1081 gatcacttgt ggactaagtt tgttcgcatc cccttctcca accccctcag tacatcaccc
     1141 tgggggaaca gggtccactt gctcctgggc ccacacagtc ctgcagtatt gtgtatataa
     1201 ggccagggca aagaggagca ggttttaaag tgaaaggcag gcaggtgttg gggaggcagt
     1261 taccggggca acgggaacag ggcgtttcgg aggtggttgc catggggacc tggatgctga
     1321 cgaaggctcg cgaggctgtg agcagccaca gtgccctgct cagaagcccc aagctcgtca
     1381 gtcaagccgg ttctccgttt gcactcagga gcacgggcag gcgagtggcc cctagttctg
     1441 ggggcagctc tagagcggta ccggatcccc gggtaccggt gccaccatgg actatggcgg
     1501 cgctttgtct gccgtcggac gcgaactttt gttcgttact aatcctgtgg tggtgaacgg
     1561 gtccgtcctg gtccctgagg atcaatgtta ctgtgccgga tggattgaat ctcgcggcac
     1621 gaacggcgct cagaccgcgt caaatgtcct gcagtggctt gcagcaggat tcagcatttt
     1681 gctgctgatg ttctatgcct accaaacctg gaaatctaca tgcggctggg aggagatcta
     1741 tgtgtgcgcc attgaaatgg ttaaggtgat tctcgagttc ttttttgagt ttaagaatcc
     1801 ctctatgctc taccttgcca caggacaccg ggtgcagtgg ctgcgctatg cagagtggct
     1861 gctcacttct cctgtcatcc ttatccacct gagcaacctc accggcctga gcaacgacta
     1921 cagcaggaga accatgggac tccttgtctc agccatcggg actatcgtgt ggggggctac
     1981 cagcgccatg gcaaccggct atgttaaagt catcttcttt tgtcttggat tgtgctatgg
     2041 cgcgaacaca ttttttcacg ccgccaaagc atatatcgag ggttatcata ctgtgccaaa
     2101 gggtcggtgc cgccaggtcg tgaccggcat ggcatggctg tttttcgtga gctggggtat
     2161 gttcccaatt ctcttcattt tggggcccga aggttttggc gtcctgagcg tctatggctc
     2221 caccgtaggt cacacgatta ttgatctgat gagtaaaaat tgttgggggt tgttgggaca
     2281 ctacctgcgc gtcctgatcc acgagcacat attgattcac ggagatatcc gcaaaaccac
     2341 caaactgaac atcggcggaa cggagatcga ggtcgagact ctcgtcgaag acgaagccga
     2401 ggccggagcc gtgccagcgg ccgccgtgag caagggcgag gaggataaca tggccatcat
     2461 caaggagttc atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga
     2521 gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt
     2581 gaccaagggt ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg
     2641 ctccaaggcc tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc
     2701 cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac
     2761 ccaggactcc tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa
     2821 cttcccctcc gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga
     2881 gcggatgtac cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa
     2941 ggacggcggc cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca
     3001 gctgcccggc gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta
     3061 caccatcgtg gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga
     3121 gctgtacaag taagaattcg atatcaagct tatcgataat caacctctgg attacaaaat
     3181 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     3241 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     3301 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     3361 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     3421 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     3481 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     3541 gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct
     3601 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     3661 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     3721 gatctccctt tgggccgcct ccccgcatcg ataccgagcg ctgctcgaga gatctacggg
     3781 tggcatccct gtgacccctc cccagtgcct ctcctggccc tggaagttgc cactccagtg
     3841 cccaccagcc ttgtcctaat aaaattaagt tgcatcattt tgtctgacta ggtgtccttc
     3901 tataatatta tggggtggag gggggtggta tggagcaagg ggcaagttgg gaagacaacc
     3961 tgtagggcct gcggggtcta ttgggaacca agctggagtg cagtggcaca atcttggctc
     4021 actgcaatct ccgcctcctg ggttcaagcg attctcctgc ctcagcctcc cgagttgttg
     4081 ggattccagg catgcatgac caggctcagc taatttttgt ttttttggta gagacggggt
     4141 ttcaccatat tggccaggct ggtctccaac tcctaatctc aggtgatcta cccaccttgg
     4201 cctcccaaat tgctgggatt acaggcgtga accactgctc ccttccctgt ccttctgatt
     4261 ttgtaggtaa ccacgtgcgg accgagcggc cgcaggaacc cctagtgatg gagttggcca
     4321 ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc
     4381 cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc
     4441 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca
     4501 accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     4561 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     4621 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     4681 ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg
     4741 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     4801 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg gctattcttt
     4861 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     4921 aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag
     4981 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     5041 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     5101 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     5161 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc
     5221 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     5281 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     5341 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     5401 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     5461 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     5521 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     5581 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     5641 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     5701 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     5761 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     5821 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     5881 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     5941 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     6001 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     6061 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     6121 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     6181 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     6241 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     6301 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     6361 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     6421 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     6481 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta
     6541 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     6601 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     6661 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     6721 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     6781 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     6841 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     6901 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     6961 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     7021 tgctcacatg t
//
