LOCUS       sequence-236913-        7046 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Zebrafish Tol1 construct for UAS-mediated expression of the 
            optogenetic opsin ChrimsonR fused to the red fluorescent protein 
            tdTomato.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7046)
  AUTHORS   Antinucci P, Dumitrescu A, Deleuze C, Morley HJ, Leung K, Hagley T, 
            Kubo F, Baier H, Bianco IH, Wyart C
  TITLE     A calibrated optogenetic toolbox of stable zebrafish opsin lines.
  JOURNAL   Elife. 2020 Mar 27;9. pii: 54937. doi: 10.7554/eLife.54937.
  PUBMED    32216873
REFERENCE   2  (bases 1 to 7046)
  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: "Elife."; 
            date: "2020-03-27"; volume: "9. pii: 54937. doi"; pages: " 
            10.7554/eLife.54937"
COMMENT     Sequence Label: pTol1-UAS:ChrimsonR-tdTomato
FEATURES             Location/Qualifiers
     source          1..7046
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(142..147,148..165,166..172)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     regulatory      1558..1567
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1564..2613
                     /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"
     primer_bind     complement(4650..4667)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4650..4666)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4659..4681)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4875..4894)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4994..5016
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5054..5072)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5140..5244
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5245..5313,5314..6105)
                     /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(5463..5482)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6276..6864
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6765..6784
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7018..7035
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
      241 atggtccctt tagccagtag cggttctagg cacgggccgt ccgggcggtg gcctggggcg
      301 gaaaactgaa ggggggcggc accggcggct cagcccttgt aatatattat atgcaccact
      361 attggtttac ttatgtcaca gtttgtaagt ttgtaacagc ctgaacctgg atcagcttag
      421 gcctccaagg cggagtactg tcctccgggc tggcggagta ctgtcctccg gcaaggtcgg
      481 agtactgtcc tccgacacta gaggtcggag tactgtcctc cgacgcaagg cggagtactg
      541 tcctccgggc tgcggagtac tgtcctccgg caaggtcgga gtactgtcct ccgacactag
      601 aggtcggagt actgtcctcc gacgcaaggt cggagtactg tcctccgaca ctagaggtcg
      661 gagtactgtc ctccgacgca aggtcggagt actgtcctcc gacactagag gtcggagtac
      721 tgtcctccga cgcaaggcgg agtactgtcc tccgggctgg cggagtactg tcctccggca
      781 agggtcgact ctagagggta tataatggat cccatcgcgt ctcagcctca ctttgagctc
      841 ctccacacga attcgatact aaggatcctt gtttgtgagt atgatgttaa atataagcaa
      901 tcgacgtttg ttttgtcgtt tgttaaatct agcaatgttt tatttcaatt aaagtgttaa
      961 ccggttcatt taaaggtgta tttctgaatg agctaacgta ttagctttta acgctagccg
     1021 gactgtttgg ttttgaggat caataaacca acgttttttt ctgatttatt ttgctttaaa
     1081 atataaaggt ataattcgtc gtcgagaatt ttgggttttg ttcgagttta gaaatttggc
     1141 ggcgtttttt tgtaccgcga aacagggcgg tggttcggtg agcttctcaa ttcatctgct
     1201 tgacaaacgg ttttcggcaa ctgtaaatgc gctatgatgc ttttattctt ttcagcgtgt
     1261 aactttgtta aatataattg actttccgac tatcgacgtt cgtttcatct ccgtgttatt
     1321 gtttaatggc cattttctgt ggtaaactgt cgccattttg ttttcagtcg tccgtttgtt
     1381 ccgcccttac tcacagctgt gtcagcatta gtgatgactg ttctcacctg gcctgtttgc
     1441 tttcattatt gatgaagagc tttctagaga attctgaagc tcgtctttgt cctgaattca
     1501 tctgtattca tttaaaaaaa aaaattaacc ttaatttttt tttcttctca gcgtaaagcc
     1561 accatggctg agctgatcag cagcgccacc agatctctgt ttgccgccgg aggcatcaac
     1621 ccttggccta acccctacca ccacgaggac atgggctgtg gaggaatgac acctacaggc
     1681 gagtgcttca gcaccgagtg gtggtgtgac ccttcttacg gactgagcga cgccggatac
     1741 ggatattgct tcgtggaggc cacaggcggc tacctggtcg tgggagtgga gaagaagcag
     1801 gcttggctgc acagcagagg cacaccagga gaaaagatcg gcgcccaggt ctgccagtgg
     1861 attgctttca gcatcgccat cgccctgctg acattctacg gcttcagcgc ctggaaggcc
     1921 acttgcggtt gggaggaggt ctacgtctgt tgcgtcgagg tgctgttcgt gaccctggag
     1981 atcttcaagg agttcagcag ccccgccaca gtgtacctgt ctaccggcaa ccacgcctat
     2041 tgcctgcgct acttcgagtg gctgctgtct tgccccgtga tcctgatcag actgagcaac
     2101 ctgagcggcc tgaagaacga ctacagcaag cggaccatgg gcctgatcgt gtcttgcgtg
     2161 ggaatgatcg tgttcggcat ggccgcagga ctggctaccg attggctcaa gtggctgctg
     2221 tatatcgtgt cttgcatcta cggcggctac atgtacttcc aggccgccaa gtgctacgtg
     2281 gaagccaacc acagcgtgcc taaaggccat tgccgcatgg tcgtgaagct gatggcctac
     2341 gcttacttcg cctcttgggg cagctaccca atcctctggg cagtgggacc agaaggactg
     2401 ctgaagctga gcccttacgc caacagcatc ggccacagca tctgcgacat catcgccaag
     2461 gagttttgga ccttcctggc ccaccacctg aggatcaaga tccacgagca catcctgatc
     2521 cacggcgaca tccggaagac caccaagatg gagatcggag gcgaggaggt ggaagtggaa
     2581 gagttcgtgg aggaggagga cgaggacaca gtggcggcac cggtagtagc agtgagtaag
     2641 ggcgaggaag tgatcaaaga gttcatgcgg tttaaggtga gaatggaagg aagcatgaac
     2701 ggccacgagt tcgaaattga gggagaagga gagggacggc cctacgaggg cacccagaca
     2761 gccaagctga aagtgacaaa gggcgggcct ctgccattcg cttgggacat cctgagccca
     2821 cagtttatgt acggctccaa ggcctatgtg aaacatccag ctgacattcc cgattataag
     2881 aaactgagct tccccgaggg gtttaagtgg gaaagagtga tgaacttcga ggacggaggc
     2941 ctggtgactg tgacccagga cagctccctg caggatggga ccctgatcta caaggtgaaa
     3001 atgagaggga caaattttcc ccctgatgga cctgtgatgc agaagaaaac tatgggatgg
     3061 gaggcctcca ccgaaaggct gtatccacgc gacggggtgc tgaaaggaga aatccaccag
     3121 gctctgaagc tgaaagatgg gggacattac ctggtggagt tcaagacaat ctacatggcc
     3181 aagaaacctg tgcagctgcc aggctactat tacgtggaca caaaactgga tatcacttca
     3241 cacaacgagg actacactat tgtggagcag tatgaacgga gcgaggggag acaccatctg
     3301 ttcctgggcc atgggactgg aagtaccggc tcagggtcta gtggaaccgc ctcaagcgag
     3361 gataacaata tggctgtgat caaagagttc atgaggttta aggtgcgcat ggagggcagc
     3421 atgaatgggc acgaatttga gattgaagga gagggcgaag ggaggcctta cgagggcaca
     3481 cagactgcca agctgaaagt gaccaaggga ggaccactgc ctttcgcttg ggatatcctg
     3541 tctcctcagt ttatgtacgg aagtaaggcc tatgtcaagc atcccgctga cattcctgat
     3601 tacaagaaac tgtctttccc agagggcttt aagtgggaga gagtgatgaa ttttgaagat
     3661 ggaggcctgg tgaccgtgac acaggactcc tctctgcagg atggcactct gatctacaaa
     3721 gtcaaaatgc gcggcaccaa ttttccaccc gatgggcccg tgatgcagaa gaaaacaatg
     3781 gggtgggagg ccagcactga acggctgtat cctagagacg gagtgctgaa gggcgaaatc
     3841 caccaggccc tgaagctgaa agacggcggc cactacctgg tggagttcaa aaccatctac
     3901 atggccaaga aaccagtgca gctgcccggc tattactatg tggacaccaa gctggatatc
     3961 acatcccaca atgaagacta caccattgtg gaacagtatg agaggtctga aggacgccac
     4021 catctgtttc tgtacggcat ggatgagctg tataagtaaa catggagatc tcgagactag
     4081 taattaagtc tcagccaccg ttaactgaac atgtcaaaac ctgtggagac tgttgagatt
     4141 tgatgttctg aaaagataaa gcctataaat aaaatgttgc ccaaatttcc tgcctgatgt
     4201 ttttctttgt ctttgctaca tggctttgct gctcggatcg gctcactctg tgtatgccac
     4261 gttcactttg tactctcctt ctcacggtag gtttattatt tttagatgtg cagttagttt
     4321 ctgtgaaata acacaccaca cactgatatt gtctgtgcat tgacttggtg agtgcacatt
     4381 gtttttgatc ttgacatatt tatatttgat tgatcaggtg aactgtgtga atctaaagtg
     4441 ctccatacag atgttctgca ttgaaaatat tctcatttta ttagtggaag tgagtgtatg
     4501 ccagcggccg cggtaccctg cagtctagac atttcagttg acgaagacaa acaaagttct
     4561 gttgtgacta tggggggggg gggcgcctgg ggatggtctc gcccggggag taattcaggg
     4621 tagaaccgcc actgccttta gcgacattca ctggccgtcg ttttacaacg tcgtgactgg
     4681 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     4741 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     4801 gaatggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     4861 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     4921 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     4981 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     5041 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     5101 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     5161 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     5221 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     5281 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     5341 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     5401 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     5461 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     5521 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     5581 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     5641 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5701 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5761 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     5821 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5881 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     5941 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     6001 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     6061 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     6121 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     6181 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6241 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6301 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6361 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6421 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6481 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6541 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6601 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6661 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6721 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6781 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6841 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6901 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     6961 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     7021 gagtcagtga gcgaggaagc ggaaga
//
