LOCUS       sequence-257676-        7652 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of ChrimsonR-tdTomato under the 
            EF1-alpha-1.1 promoter in frt/reversed (Flp-dependent) manner. 
            tdTomato has codons varied to reduce recombination..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7652)
  AUTHORS   Klapoetke NC, Murata Y, Kim SS, Pulver SR, Birdsey-Benson A, Cho YK,
            Morimoto TK, Chuong AS, Carpenter EJ, Tian Z, Wang J, Xie Y, Yan Z, 
            Zhang Y, Chow BY, Surek B, Melkonian M, Jayaraman V, 
            Constantine-Paton M, Wong GK, Boyden ES
  TITLE     Independent optical excitation of distinct neural populations.
  JOURNAL   Nat Methods. 2014 Mar;11(3):338-46. doi: 10.1038/nmeth.2836. Epub 
            2014 Feb 9.
  PUBMED    24509633
REFERENCE   2  (bases 1 to 7652)
  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"; doi: 
            "10.1038/nmeth.2836"; journalName: "Nat Methods"; date: "2014-03"; 
            volume: "11"; issue: "3"; pages: "338-46"
COMMENT     Sequence Label: AAV-EF1-alpha-1.1-FLPX-rc [ChrimsonR-tdTomato]
FEATURES             Location/Qualifiers
     source          1..7652
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        145..1323
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          376..1314
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1271..1291
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    1330..1363
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             complement(2903..3952)
                     /codon_start=1
                     /product="red light-drivable channelrhodopsin from 
                     Chlamydomonas noctigama (Klapoetke et al., 2014)"
                     /label=ChrimsonR
                     /note="K176R mutant with accelerated closing kinetics"
                     /translation="MAELISSATRSLFAAGGINPWPNPYHHEDMGCGGMTPTGECFSTE
                     WWCDPSYGLSDAGYGYCFVEATGGYLVVGVEKKQAWLHSRGTPGEKIGAQVCQWIAFSI
                     AIALLTFYGFSAWKATCGWEEVYVCCVEVLFVTLEIFKEFSSPATVYLSTGNHAYCLRY
                     FEWLLSCPVILIRLSNLSGLKNDYSKRTMGLIVSCVGMIVFGMAAGLATDWLKWLLYIV
                     SCIYGGYMYFQAAKCYVEANHSVPKGHCRMVVKLMAYAYFASWGSYPILWAVGPEGLLK
                     LSPYANSIGHSICDIIAKEFWTFLAHHLRIKIHEHILIHGDIRKTTKMEIGGEEVEVEE
                     FVEEEDEDTV"
     regulatory      3949..3958
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3976..4009)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    4099..4687
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4152..4172)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4570..4581)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     repeat_region   4915..5055
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4915..5044
                     /label=AAV2 ITR
     rep_origin      5130..5585
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5217..5236)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5427..5448
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5602..5621)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5721..5743
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5781..5799)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5867..5971
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5972..6040,6041..6832)
                     /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(6190..6209)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7003..7591
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7492..7511
                     /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 gtcgacttaa ttaaggctcc ggtgcccgtc agtgggcaga gcgcacatcg
      181 cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gaaccggtgc ctagagaagg
      241 tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt tcccgagggt
      301 gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg caacgggttt
      361 gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg ggcctggcct ctttacgggt
      421 tatggccctt gcgtgccttg aattacttcc acctggctgc agtacgtgat tcttgatccc
      481 gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt gcgcttaagg agccccttcg
      541 cctcgtgctt gagttgaggc ctggcctggg cgctggggcc gccgcgtgcg aatctggtgg
      601 caccttcgcg cctgtctcgc tgctttcgat aagtctctag ccatttaaaa tttttgatga
      661 cctgctgcga cgcttttttt ctggcaagat agtcttgtaa atgcgggcca agatctgcac
      721 actggtattt cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc ccagcgcaca
      781 tgttcgacga ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg gtagtctcaa
      841 gctggccggc ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc gccctgggcg
      901 gcaaggctgg cccggtcggc accagttgcg tgagcggaaa gatggccgct tcccggccct
      961 gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg tgagtcaccc
     1021 acacaaagga aaagggcctt tccgtcctca gccgtcgctt catgtgactc cacggagtac
     1081 cgggcgccgt ccaggcacct cgattagttc tcgagctttt ggagtacgtc gtctttaggt
     1141 tggggggagg ggttttatgc gatggagttt ccccacactg agtgggtgga gactgaagtt
     1201 aggccagctt ggcacttgat gtaattctcc ttggaatttg ccctttttga gtttggatct
     1261 tggttcattc tcaagcctca gacagtggtt caaagttttt ttcttccatt tcaggtgtcg
     1321 tgagctagcg aagttcctat tctctagaaa gtataggaac ttcgcagaat ggtagctgga
     1381 ttgtagctgc tattagcaat atgaaacctc ttagaagttc ctattcttca aaaggtatag
     1441 gaacttcgaa ttctcattac ttatacagct catccatgcc gtacagaaac agatggtggc
     1501 gtccttcaga cctctcatac tgttccacaa tggtgtagtc ttcattgtgg gatgtgatat
     1561 ccagcttggt gtccacatag taatagccgg gcagctgcac tggtttcttg gccatgtaga
     1621 tggttttgaa ctccaccagg tagtggccgc cgtctttcag cttcagggcc tggtggattt
     1681 cgcccttcag cactccgtct ctaggataca gccgttcagt gctggcctcc caccccattg
     1741 ttttcttctg catcacgggc ccatcgggtg gaaaattggt gccgcgcatt ttgactttgt
     1801 agatcagagt gccatcctgc agagaggagt cctgtgtcac ggtcaccagg cctccatctt
     1861 caaaattcat cactctctcc cacttaaagc cctctgggaa agacagtttc ttgtaatcag
     1921 gaatgtcagc gggatgcttg acataggcct tacttccgta cataaactga ggagacagga
     1981 tatcccaagc gaaaggcagt ggtcctccct tggtcacttt cagcttggca gtctgtgtgc
     2041 cctcgtaagg cctcccttcg ccctctcctt caatctcaaa ttcgtgccca ttcatgctgc
     2101 cctccatgcg caccttaaac ctcatgaact ctttgatcac agccatattg ttatcctcgc
     2161 ttgaggcggt tccactagac cctgagccgg tacttccagt cccatggccc aggaacagat
     2221 ggtgtctccc ctcgctccgt tcatactgct ccacaatagt gtagtcctcg ttgtgtgaag
     2281 tgatatccag ttttgtgtcc acgtaatagt agcctggcag ctgcacaggt ttcttggcca
     2341 tgtagattgt cttgaactcc accaggtaat gtcccccatc tttcagcttc agagcctggt
     2401 ggatttctcc tttcagcacc ccgtcgcgtg gatacagcct ttcggtggag gcctcccatc
     2461 ccatagtttt cttctgcatc acaggtccat cagggggaaa atttgtccct ctcattttca
     2521 ccttgtagat cagggtccca tcctgcaggg agctgtcctg ggtcacagtc accaggcctc
     2581 cgtcctcgaa gttcatcact ctttcccact taaacccctc ggggaagctc agtttcttat
     2641 aatcgggaat gtcagctgga tgtttcacat aggccttgga gccgtacata aactgtgggc
     2701 tcaggatgtc ccaagcgaat ggcagaggcc cgccctttgt cactttcagc ttggctgtct
     2761 gggtgccctc gtagggccgt ccctctcctt ctccctcaat ttcgaactcg tggccgttca
     2821 tgcttccttc cattctcacc ttaaaccgca tgaactcttt gatcacttcc tcgcccttac
     2881 tcactgctac taccggtgcc gccactgtgt cctcgtcctc ctcctccacg aactcttcca
     2941 cttccacctc ctcgcctccg atctccatct tggtggtctt ccggatgtcg ccgtggatca
     3001 ggatgtgctc gtggatcttg atcctcaggt ggtgggccag gaaggtccaa aactccttgg
     3061 cgatgatgtc gcagatgctg tggccgatgc tgttggcgta agggctcagc ttcagcagtc
     3121 cttctggtcc cactgcccag aggattgggt agctgcccca agaggcgaag taagcgtagg
     3181 ccatcagctt cacgaccatg cggcaatggc ctttaggcac gctgtggttg gcttccacgt
     3241 agcacttggc ggcctggaag tacatgtagc cgccgtagat gcaagacacg atatacagca
     3301 gccacttgag ccaatcggta gccagtcctg cggccatgcc gaacacgatc attcccacgc
     3361 aagacacgat caggcccatg gtccgcttgc tgtagtcgtt cttcaggccg ctcaggttgc
     3421 tcagtctgat caggatcacg gggcaagaca gcagccactc gaagtagcgc aggcaatagg
     3481 cgtggttgcc ggtagacagg tacactgtgg cggggctgct gaactccttg aagatctcca
     3541 gggtcacgaa cagcacctcg acgcaacaga cgtagacctc ctcccaaccg caagtggcct
     3601 tccaggcgct gaagccgtag aatgtcagca gggcgatggc gatgctgaaa gcaatccact
     3661 ggcagacctg ggcgccgatc ttttctcctg gtgtgcctct gctgtgcagc caagcctgct
     3721 tcttctccac tcccacgacc aggtagccgc ctgtggcctc cacgaagcaa tatccgtatc
     3781 cggcgtcgct cagtccgtaa gaagggtcac accaccactc ggtgctgaag cactcgcctg
     3841 taggtgtcat tcctccacag cccatgtcct cgtggtggta ggggttaggc caagggttga
     3901 tgcctccggc ggcaaacaga gatctggtgg cgctgctgat cagctcagcc atggtggcgg
     3961 atccggcggc gcgccgaagt tcctatactt tctagagaat aggaacttct tgccttaacc
     4021 cagaaattat cactgttatt ctttagaatg gtgcaaagag aagttcctat accttttgaa
     4081 gaataggaac ttctccggaa tcaacctctg gattacaaaa tttgtgaaag attgactggt
     4141 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat
     4201 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg
     4261 tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt
     4321 gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact
     4381 ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
     4441 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaagctgacg
     4501 tcctttccat ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc
     4561 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg
     4621 cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc
     4681 tccccgcctt gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     4741 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     4801 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     4861 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcaggaac
     4921 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
     4981 gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc
     5041 gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     5101 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
     5161 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc
     5221 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     5281 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     5341 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     5401 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     5461 ctctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt
     5521 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     5581 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     5641 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     5701 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     5761 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     5821 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     5881 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     5941 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     6001 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     6061 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     6121 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     6181 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     6241 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     6301 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     6361 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     6421 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     6481 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     6541 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     6601 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     6661 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     6721 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     6781 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     6841 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     6901 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     6961 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     7021 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     7081 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     7141 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     7201 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     7261 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     7321 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     7381 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     7441 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     7501 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     7561 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     7621 cctggccttt tgctggcctt ttgctcacat gt
//
