LOCUS       sequence-206463-        9528 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Muscle specific zebrafish expression of mKate2 fused to dominant 
            negative RhoAa.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9528)
  AUTHORS   Hall TE, Martel N, Ariotti N, Xiong Z, Lo HP, Ferguson C, Rae J, Lim
            YW, Parton RG
  TITLE     In vivo cell biological screening identifies an endocytic capture 
            mechanism for T-tubule formation.
  JOURNAL   Nat Commun. 2020 Jul 24;11(1):3711. doi: 10.1038/s41467-020-17486-w.
  PUBMED    32709891
REFERENCE   2  (bases 1 to 9528)
  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: "Nat 
            Commun."; date: "2020-07-24"; volume: "11(1):3711. doi"; pages: " 
            10.1038/s41467-020-17486-w"
COMMENT     Sequence Label: actc1b-mKate2-RhoAa-DN
FEATURES             Location/Qualifiers
     source          1..9528
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(144..165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(514..525,526..688)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     primer_bind     1061..1077
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1097..1117
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    5092..5116
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      5117..5126
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5123..5818
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     protein_bind    complement(5820..5844)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(6425..6445)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     complement(6453..6469)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    6485..6619
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6509..6528)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6563..6582
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(7058..7245,7246..7257)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(7291..7307)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7291..7307)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(7344..7364)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(7344..7362)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7383..7399)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7396..7418)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7407..7423
                     /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)."
     promoter        complement(join(7431..7437,7438..7455,7456..7461))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7476..7497
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7614..7631)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7785..8373)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7865..7884)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8544..9335,9336..9404))
                     /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     9167..9186
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9405..9509)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
       61 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
      121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
      181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacgc tggctaagaa
      241 ctcatcagcc tccccggtcc atctacccac gtaccaatgc accaattggc cacaatgacg
      301 gctactacat ggtgccattc cttcctcttt ataggaatgg agactacctc ctgtccaaca
      361 aggctcttgg atacgagtac gcctacctgt tggacccagg tcattgcaca acaccagaaa
      421 tgccctctga tctgcaaaag acgtgaatat ctgttcagac acccatatcc actctgttcc
      481 acacaggtca gaggtttgtc caggagttct tgacagaggt gtaaaaagta ctcaaaaatt
      541 ttactcaagt gaaagtacaa gtacttaggg aaaattttac tcaattaaaa gtaaaagtat
      601 ctggctagaa tcttacttga gtaaaagtaa aaaagtactc cattaaaatt gtacttgagt
      661 attaaggaag taaaagtaaa agcaagaaag aaaactagag attcttgttt aagcttttaa
      721 tctcaaaaaa cattaaatga aatgcataca aggttttatc ctgctttaga actgtttgta
      781 tttaattatc aaactataag acagacaatc taatgccagt acacgctact caaagttgta
      841 aaacctcaga tttaacttca gtagaagctg attctcaaaa ttgttagtgt caagcctagc
      901 tcttttgggg ctgaaaagca atcctgcagt gctgaaaagc ctctcacagg cagccgatgc
      961 gggaagaggt gtattagtct tgatagagag gctgcaaata gcaggaaacg tgagcagaga
     1021 ctccctggtg tctgaaacac aggccagatg ggccctcgag caggaaacag ctatgaccat
     1081 gattacgcca agctatcaac tttgtataga aaagttggca cgcgcagatt ccgtgtgggc
     1141 ctggtaatat cttaccattt taagctattt taagattagg gaaaaaaatt aaataaaact
     1201 actttaaaat tactttcctt aaaaagcaac atatttattt agtttttgtt taactggctc
     1261 aagattgtaa tattttactt ttacatgttt tttttaagta cctaaatgtg ttcaaaagta
     1321 aagtatgtta ttgttttttt ttttttaatg ttagctaagc tttaaattgt aatatattac
     1381 ttttaaaagt gactttttaa aaatgtaaat aaaaaagtaa ctataaatgt gtaatgcatg
     1441 ttatttctgt gggtttttta attttcatct caactgggct tacaatttta acataataaa
     1501 aatcaaatca aatcaaatca ggcccttatt tgtcacatac actcttgtat actgaaataa
     1561 actaaatgtt gagaaaaatt taaatacacc aaaataacgc ataccttttt tgtaattacc
     1621 taagtaaaac atttagttcc ttttttaagt gactcaagat tgtaatctat taattgtaaa
     1681 agataaaact agtgatcaat ctgacgtgct tctatgcagc ctgtgagatt aattgtgatg
     1741 tttaattatt aaacaattta aatggatcaa acatttcaca tgtgcttatt gtgtccaaca
     1801 aaaccagcag aagcacttaa aaatattgca tgacaacatg agtttttaat gtaaattata
     1861 tatacttcct aaacgaagtt aatcatattt tctaaaataa taatgtcatt gaaatattta
     1921 aataaatgat aanctttagt tgattcaaca taaaattgaa agcaataagg actttttttt
     1981 ttttacagcg gtgaagcaaa agcntgnaaa taatcatata aatataataa ataaataaat
     2041 aaaataatgg ttcttggtgc aaatatgaac actttaaata aatatagaat aaatatgatc
     2101 aataagtcat aacgacgtgt tttatgttga ttataattta aacatgtttc taacgcaagt
     2161 tttaaaaact gttttaaagt attattttaa gttataagat caatttgact caacattttg
     2221 tagtgaaaat atgttgtgta aaattaatat aatatacact ataaaaatgc aaggttccac
     2281 acaattcatt catgttgtcc caacacaaat cgattaagtt aacaaatata tgtggattga
     2341 acataaaaaa ttaagttgtc ccaatgaaat ctcaagaatt gtgttgtttc agctcatttt
     2401 aaataagtag ttttaacaag taaaaaataa catatttttg agtgtgtata tatatatatt
     2461 tatatattac aaaatataat aatataaaat gtaaaagata taaaccatgc tacatatatt
     2521 taagagctat tttgcacatt acttttttgc aagccatttg agtgtttttt tacaaatata
     2581 tattagctaa atatacaata tcttaaatat acaattaatc agacatgcaa aaaatgcatt
     2641 taacctacat aaaaatacat gatcaacata tgttttatgc tgattacaac taaatcaaac
     2701 gtaatgaaat taatttataa aagcgattta aatatgtcaa tattttaagt taaaatcact
     2761 tgaaacctac tataaatgaa cttaaattta aaaaatctcc tttaattcca atgcagcact
     2821 gttgtttttt cagctgagga cccccggtgc agatggttgg cccccggccc gctattctaa
     2881 ggaagccttt cttaatttta gggcggcgct gcatgcgctc agatcagatt cacgggtgga
     2941 gagggttatt ttctcttttt tattttttct tgcctgctca tatttgggcc tgtgtgaacc
     3001 ccaggctccg cctcggcccc caggagccgc tcttatttaa aggggatcct cgccttcagt
     3061 cttcgtccca gtggaagcat cagtcttgtg ataacaactt ggtgagtcct acagaccagc
     3121 gctcgactct aggctttaaa tgcattgatt tctctggatt agctaccggg agagttttaa
     3181 ctttgcttat tagtcttgtg cttgaactgc taggtttaaa gagggttttc tctaatgcat
     3241 gcatgaagcg acagctgaaa accagaaata gagcgtttgt cctcctgtta aactctagcc
     3301 tttctttggc atttgtcagt ccgagtcttc agctgccggc aggagattgg gtttcagtaa
     3361 cacttgctga gcacaaacta cacgtgcttc cttcagattt aattcattat taatcattct
     3421 agatttaaat gatcttctta ctcattgctg attattttag ccttgcggag cagattaaat
     3481 gagcacgtgt ttattaagat tgatttaaat ataattaata tttcaccact gtagttttaa
     3541 aattgccttc taagtcatgg ttgattttta ataggtatcg gctctaatta gattactatt
     3601 ttggttttta aatgttattt ctatttcatt tgtaggctcg tttttaatgt gattttatta
     3661 gattctagtt ttaaaatcat cttttagcct tactgagcag actgaatgaa catgtttcat
     3721 taagattaac ttcattatag ttaattattt aatcattcta gtggattata aatcagctct
     3781 atttaaattg attgtcgtgt ttatttctat ttgatttttg attatttttt aaggtcgttt
     3841 ttatgtgatt tttccccgat tctaatttta aactaattgt ttgattttta tatgattcga
     3901 ctctttttaa atgaattgtt ttgctttttt tattgagcat atatatatat tatacaattt
     3961 caatcaacaa ttgttgtgct ttttaaatgt tgtttctatt tcatttctag ggtcgatttt
     4021 attcgattct agttttaaaa tcgtgatttc taactcctaa ccttactgag cagactgaat
     4081 gaacatgttt catcaagatt aacttcatta tagttaatta ttgaatcggg ctagtggttt
     4141 ataaatcagc tctattataa attaattgtc atcattattt ctatttaatt tttattattt
     4201 ttttaggatt atttttaagg gatttttttc acgttctagt tttaaactca atgctcatta
     4261 ctgctcaccc ttgctgagca gtctttcgtt aagattaaat taattaataa cccattctag
     4321 ttttaaaaca gtgtcttaag gcatagctga tattttacaa gtttactatt taatatatct
     4381 ccatttcttt ttttagggtc gatttcatct gatttgatta atttttatta tcttcactaa
     4441 gctttttgtt ctaggtttta ttctcagtat tattactgtt atatttgaac tgaatttctc
     4501 ctcatagttt attcatttta tttatttatt ttttgtttat tcgctatttt gtattttagt
     4561 ttttaattag aaattttatt agaatcgttt tgaactagtt tttatgcact ctaacattaa
     4621 tattaaatca taactattat ttttattata atttatttgt gtttatcttt ttgtgtagct
     4681 ttcatgtgtt tttattttag ttttattatg tttgcattga attttttgga ctagtttata
     4741 taacgatata cagaaaagcc atgacaacaa atgtttttat aatacctatt aagtataagc
     4801 taatcagatt tccactattc tgttcaaagt taacattatt agccattctg atggatcatt
     4861 tgacatttgt catatgtctt tatgaacact gctctttcag atgccacttt taaagctgtt
     4921 ttttacagta aatgaacgtt tattattatg acattatttc ctaaggcctc tagagtacag
     4981 caggatgtgg cgtgtgcatt actgtgtaat tatgctgact gtaaatgtgc atttatagac
     5041 tgtcagcatg tctgactgtg cgctgttgtg atttgtcctg cacagaccaa ggcaagtttg
     5101 tacaaaaaag caggctgcca ccatggtgag cgagctgatt aaggagaaca tgcacatgaa
     5161 gctgtacatg gagggcaccg tgaacaacca ccacttcaag tgcacatccg agggcgaagg
     5221 caagccctac gagggcaccc agaccatgag aatcaaggcg gtcgagggcg gccctctccc
     5281 cttcgccttc gacatcctgg ctaccagctt catgtacggc agcaaaacct tcatcaacca
     5341 cacccagggc atccccgact tctttaagca gtccttcccc gagggcttca catgggagag
     5401 agtcaccaca tacgaagacg ggggcgtgct gaccgctacc caggacacca gcctccagga
     5461 cggctgcctc atctacaacg tcaagatcag aggggtgaac ttcccatcca acggccctgt
     5521 gatgcagaag aaaacactcg gctgggaggc ctccaccgag accctgtacc ccgctgacgg
     5581 cggcctggaa ggcagagccg acatggccct gaagctcgtg ggcgggggcc acctgatctg
     5641 caacttgaag accacataca gatccaagaa acccgctaag aacctcaaga tgcccggcgt
     5701 ctactatgtg gacagaagac tggaaagaat caaggaggcc gacaaagaga cctacgtcga
     5761 gcagcacgag gtggctgtgg ccagatactg cgacctccct agcaaactgg ggcacagaga
     5821 cccagctttc ttgtacaaag tggtggctgc gattcgtaag aagcttgtaa tcgttggaga
     5881 tgtagcctgt ggaaagacgt gtttactcat cgttttcagt aaagaccagt ttcctgaggt
     5941 ttacgttccc acagtgtttg agaactatgt cgctgatatt gaggtggatt caaaacaggt
     6001 ggaactggct ctatgggata cagctggcca agaggactac gacaggctga ggcccctgtc
     6061 ctacccagac actgatgtca tccttatgtg cttttccata gacagtcctg acagtttgga
     6121 gaacatccca gagaaatgga cgccagaagt aaaacatttc tgccctaatg ttccaataat
     6181 actggtgggt aataaaaaag atttacggaa tgacgagcac actcgacggg agctgcagaa
     6241 gatgaaacag gaaccagtta aaccagagga gggacgagac atggctaatc gcatcaatgc
     6301 cttcggctac cttgagtgtt cagctaaaac taaggagggc gtgagggaag tgtttgaaat
     6361 ggccacaaga gcggcgctcc aagccaagaa acgtggcaag aagaatgcct gcgctttgct
     6421 atagcaactt tattatacat agttgataat tcactggccg tcgttttacg gtaccatcga
     6481 tgatgatcca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     6541 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     6601 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     6661 ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta tggctgatta
     6721 tgatcctcta gatcagatct gcgaagatac ggccacgggt gctcttgatc ctgtggctga
     6781 ttttggactg tgctgctcgc agctgctgat gaatcacata cttcctccat tttcttccac
     6841 tgattgactg ttataatttc cctaatttcc aggtcaaggt gctgtgcatt gtggtaatag
     6901 atgtgacatg acgtcacttc caaaggacca atgaacatgt ctgaccaatt tcatataatg
     6961 tgaaaacgat tttcataggc agaataaata acatttaaat taaactgggc atcagcgcaa
     7021 ttcaattggt ttggtaatag caagggaaaa tagaatgaag tgatctccaa aaaataagta
     7081 ctttttgact gtaaataaaa ttgtaaggag taaaaagtac ttttttttct aaaaaaatgt
     7141 aattaagtaa aagtaaaagt attgattttt aattgtactc aagtaaagta aaaatcccca
     7201 aaaataatac ttaagtacag taatcaagta aaattactca agtactttac acctctggtt
     7261 cttgaccccc taccttcagc aagcccagca gatccactag ttctagagcg gccgccaccg
     7321 cggtggagct ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
     7381 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     7441 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     7501 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     7561 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     7621 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     7681 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     7741 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     7801 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     7861 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     7921 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     7981 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     8041 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     8101 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     8161 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     8221 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     8281 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     8341 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     8401 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     8461 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     8521 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     8581 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     8641 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     8701 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     8761 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     8821 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     8881 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     8941 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     9001 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     9061 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     9121 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     9181 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     9241 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     9301 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     9361 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     9421 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     9481 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtg
//
