LOCUS       Exported                6915 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV production of QuasAr2 fluorescent reporter.
ACCESSION   .
VERSION     .
KEYWORDS    DRH322: AAV-CaMKIIa-QuasAr2-mO2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6915)
  AUTHORS   Hochbaum DR, Zhao Y, Farhi SL, Klapoetke N, Werley CA, Kapoor V, Zou
            P, Kralj JM, Maclaurin D, Smedemark-Margulies N, Saulnier JL, 
            Boulting GL, Straub C, Cho YK, Melkonian M, Wong GK, Harrison DJ, 
            Murthy VN, Sabatini BL, Boyden ES, Campbell RE, Cohen AE
  TITLE     All-optical electrophysiology in mammalian neurons using engineered 
            microbial rhodopsins.
  JOURNAL   Nat Methods. 2014 Aug;11(8):825-33. doi: 10.1038/nmeth.3000. Epub 
            2014 Jun 22.
  PUBMED    24952910
REFERENCE   2  (bases 1 to 6915)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6915
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        149..1437
                     /label=CaMKII promoter
                     /note="promoter for the mouse calcium/calmodulin-dependent 
                     kinase II alpha subunit"
     CDS             1461..2219
                     /codon_start=1
                     /product="achaerhodopsin-3 (Arch) derivative with improved 
                     voltage sensitivity (Hochbaum et al., 2014)"
                     /label=QuasAr2
                     /translation="MVSIALQAGYDLLGDGRPETLWLGIGTLLMLIGTFYFLVRGWGVT
                     DKDAREYYAVTILVSGIASAAYLSMFFGIGLTEVSVGGEMLDIYYARYAQWLFTTPLLL
                     LHLALLAKVDRVTIGTLVGVDALMIVTGLIGALSHTAIARYSWWLFSTICMIVVLYVLA
                     TSLRSAAKERGPEVASTFNTLTALVLVLWTAYPILWIIGTEGAGVVGLGIETLLFMVLD
                     VTAKVGFGFILLRSRAILGDTEAPEPSAGAD"
     CDS             2226..2933
                     /codon_start=1
                     /product="photostable monomeric orange derivative of DsRed 
                     fluorescent protein (Shaner et al., 2008)"
                     /label=mOrange2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEYNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     FQTAKLKVTKGGPLPFAWDILSPHFTYGSKAYVKHPADIPDYFKLSFPEGFKWERVMNY
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GKIKMRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     misc_feature    3042..3630
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3095..3115)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3513..3524)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3662..4138
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3861..3880)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4178..4318
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4178..4307
                     /label=AAV2 ITR
     rep_origin      4393..4848
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4480..4499)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4690..4711
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4865..4884)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4984..5006
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5044..5062)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5130..5234
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5235..6095
                     /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(5453..5472)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6266..6854
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6755..6774
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtttaaca ttatggcctt aggtcacttc atctccatgg
      181 ggttcttctt ctgattttct agaaaatgag atgggggtgc agagagcttc ctcagtgacc
      241 tgcccagggt cacatcagaa atgtcagagc tagaacttga actcagatta ctaatcttaa
      301 attccatgcc ttgggggcat gcaagtacga tatacagaag gagtgaactc attagggcag
      361 atgaccaatg agtttaggaa agaagagtcc agggcagggt acatctacac cacccgccca
      421 gccctgggtg agtccagcca cgttcacctc attatagttg cctctctcca gtcctacctt
      481 gacgggaagc acaagcagaa actgggacag gagccccagg agaccaaatc ttcatggtcc
      541 ctctgggagg atgggtgggg agagctgtgg cagaggcctc aggaggggcc ctgctgctca
      601 gtggtgacag ataggggtga gaaagcagac agagtcattc cgtcagcatt ctgggtctgt
      661 ttggtacttc ttctcacgct aaggtggcgg tgtgatatgc acaatggcta aaaagcaggg
      721 agagctggaa agaaacaagg acagagacag aggccaagtc aaccagacca attcccagag
      781 gaagcaaaga aaccattaca gagactacaa gggggaaggg aaggagagat gaattagctt
      841 cccctgtaaa ccttagaacc cagctgttgc cagggcaacg gggcaatacc tgtctcttca
      901 gaggagatga agttgccagg gtaactacat cctgtctttc tcaaggacca tcccagaatg
      961 tggcacccac tagccgttac catagcaact gcctctttgc cccacttaat cccatcccgt
     1021 ctgttaaaag ggccctatag ttggaggtgg gggaggtagg aagagcgatg atcacttgtg
     1081 gactaagttt gttcgcatcc ccttctccaa ccccctcagt acatcaccct gggggaacag
     1141 ggtccacttg ctcctgggcc cacacagtcc tgcagtattg tgtatataag gccagggcaa
     1201 agaggagcag gttttaaagt gaaaggcagg caggtgttgg ggaggcagtt accggggcaa
     1261 cgggaacagg gcgtttcgga ggtggttgcc atggggacct ggatgctgac gaaggctcgc
     1321 gaggctgtga gcagccacag tgccctgctc agaagcccca agctcgtcag tcaagccggt
     1381 tctccgtttg cactcaggag cacgggcagg cgagtggccc ctagttctgg gggcagctct
     1441 agagcggtac cggatccacc atggtaagta tcgctctgca ggctggttac gacctactgg
     1501 gtgacggcag acctgaaacc ctgtggctgg gcatcggcac tctgctgatg ctgattggaa
     1561 ccttctactt tctggtccgc ggatggggag tcaccgataa ggatgcccgg gaatattacg
     1621 ctgtgactat cctggtgtcc ggaatcgcat ccgccgcata tctgtctatg ttctttggta
     1681 tcgggcttac tgaggtgtcc gtcgggggcg aaatgttgga tatctattat gccaggtacg
     1741 cccagtggct gtttaccacc ccacttctgc tgctgcacct ggcccttctc gctaaggtgg
     1801 atcgggtgac catcggcacc ctggtgggtg tggacgccct gatgatcgtc actggcctca
     1861 tcggagcctt gagccacacg gccatagcca gatacagttg gtggttgttc tctacaattt
     1921 gcatgatagt ggtgctctat gttctggcta catccctgcg atctgctgca aaggagcggg
     1981 gccccgaggt ggcatctacc tttaacaccc tgacagctct ggtcttggtg ctgtggaccg
     2041 cttaccctat cctgtggatc ataggcactg agggcgctgg cgtggtgggc ctgggcatcg
     2101 aaactctgct gtttatggtg ttggacgtga ctgccaaggt cggctttggc tttatcctgt
     2161 tgagatcccg ggctattctg ggcgacaccg aggcaccaga acccagtgcc ggtgccgata
     2221 cgttgatggt gagcaagggc gaggagtata acatggccat catcaaggag ttcatgcgct
     2281 tcaaggtgcg catggagggc tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg
     2341 agggccgccc ctacgagggc tttcagaccg ctaagctgaa ggtgaccaag ggtggccccc
     2401 tgcccttcgc ctgggacatc ctgtcccctc atttcaccta cggctccaag gcctacgtga
     2461 agcaccccgc cgacatcccc gactacttca agctgtcctt ccccgagggc ttcaagtggg
     2521 agcgcgtgat gaactacgag gacggcggcg tggtgaccgt gacccaggac tcctccctgc
     2581 aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc tccgacggcc
     2641 ccgtgatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg taccccgagg
     2701 acggtgccct gaagggcaag atcaagatga ggctgaagct gaaggacggc ggccactaca
     2761 cctccgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc ggcgcctaca
     2821 tcgtcgacat caagttggac atcacctccc acaacgagga ctacaccatc gtggaacagt
     2881 acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac aagagtagaa
     2941 tcacaagcga aggcgagtac atccccctgg atcaaataga cataaatgta ggtggatttt
     3001 gttatgagaa tgaataagaa ttcgatatca agcttatcga taatcaacct ctggattaca
     3061 aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat
     3121 acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct
     3181 ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac
     3241 gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca
     3301 cctgtcagct cctttccgga actttcgctt tccccctccc tattgccacg gcggaactca
     3361 tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg
     3421 tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctatgtt gccacctgga
     3481 ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt
     3541 cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga
     3601 gtcggatctc cctttgggcc gcctccccgc atcgataccg agcgctgctc gagagatcta
     3661 cgggtggcat ccctgtgacc cctccccagt gcctctcctg gccctggaag ttgccactcc
     3721 agtgcccacc agccttgtcc taataaaatt aagttgcatc attttgtctg actaggtgtc
     3781 cttctataat attatggggt ggaggggggt ggtatggagc aaggggcaag ttgggaagac
     3841 aacctgtagg gcctgcgggg tctattggga accaagctgg agtgcagtgg cacaatcttg
     3901 gctcactgca atctccgcct cctgggttca agcgattctc ctgcctcagc ctcccgagtt
     3961 gttgggattc caggcatgca tgaccaggct cagctaattt ttgttttttt ggtagagacg
     4021 gggtttcacc atattggcca ggctggtctc caactcctaa tctcaggtga tctacccacc
     4081 ttggcctccc aaattgctgg gattacaggc gtgaaccact gctcccttcc ctgtccttct
     4141 gattttgtag gtaaccacgt gcggaccgag cggccgcagg aacccctagt gatggagttg
     4201 gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga
     4261 cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg
     4321 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa
     4381 agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     4441 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     4501 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     4561 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt
     4621 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     4681 tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt
     4741 cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt
     4801 aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc
     4861 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
     4921 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     4981 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa
     5041 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     5101 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     5161 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5221 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5281 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     5341 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     5401 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     5461 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     5521 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     5581 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     5641 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     5701 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     5761 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     5821 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     5881 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     5941 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     6001 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     6061 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     6121 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     6181 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     6241 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6301 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     6361 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     6421 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     6481 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     6541 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     6601 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     6661 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     6721 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     6781 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     6841 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     6901 cttttgctca catgt
//
