LOCUS       sequence-224852-        9456 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Muscle specific zebrafish expression of mKate2 fused to dominant 
            negative ARF6.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9456)
  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 9456)
  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-ARF6-DN
FEATURES             Location/Qualifiers
     source          1..9456
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..25
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      26..35
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             32..727
                     /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(729..753)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1280..1300)
                     /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(1308..1324)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    1340..1474
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1364..1383)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1418..1437
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(1913..2100,2101..2112)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(2146..2162)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2146..2162)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2199..2219)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2199..2217)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2238..2254)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2251..2273)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2262..2278
                     /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(2286..2292,2293..2310,2311..2316))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2331..2352
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2469..2486)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2640..3228)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2720..2739)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3399..4190,4191..4259))
                     /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     4022..4041
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4260..4364)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4527..4548)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(4897..4908,4909..5071)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     primer_bind     5444..5460
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5480..5500
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
ORIGIN
        1 gcaagtttgt acaaaaaagc aggctgccac catggtgagc gagctgatta aggagaacat
       61 gcacatgaag ctgtacatgg agggcaccgt gaacaaccac cacttcaagt gcacatccga
      121 gggcgaaggc aagccctacg agggcaccca gaccatgaga atcaaggcgg tcgagggcgg
      181 ccctctcccc ttcgccttcg acatcctggc taccagcttc atgtacggca gcaaaacctt
      241 catcaaccac acccagggca tccccgactt ctttaagcag tccttccccg agggcttcac
      301 atgggagaga gtcaccacat acgaagacgg gggcgtgctg accgctaccc aggacaccag
      361 cctccaggac ggctgcctca tctacaacgt caagatcaga ggggtgaact tcccatccaa
      421 cggccctgtg atgcagaaga aaacactcgg ctgggaggcc tccaccgaga ccctgtaccc
      481 cgctgacggc ggcctggaag gcagagccga catggccctg aagctcgtgg gcgggggcca
      541 cctgatctgc aacttgaaga ccacatacag atccaagaaa cccgctaaga acctcaagat
      601 gcccggcgtc tactatgtgg acagaagact ggaaagaatc aaggaggccg acaaagagac
      661 ctacgtcgag cagcacgagg tggctgtggc cagatactgc gacctcccta gcaaactggg
      721 gcacagagac ccagctttct tgtacaaagt ggctgggaag atgctttcca agatcttcgg
      781 caacaaggag atgagaatat tgatgctcgg acttgacgcc gccggaaaga acacgattct
      841 gtacaaactc aaactgggcc agtcggttac gaccatcccg acagtgggct tcaatgtaga
      901 gactgtcacg tataaaaacg tcaagtttaa cgtttgggac gtcggcggtc aggataagat
      961 ccgtcctcta tggcggcact actacaccgg cacgcagggc ctgattttcg tagtggactg
     1021 tgcggatcgt gaccgcatcg atgaggctcg gcaggagctg caccgcatta tcaacgatcg
     1081 tgagatgcgc gacgccatta tcctgatctt cgccaacaaa caagaccttc cagacgccat
     1141 gaagccgcat gagatacagg agaagctggg cctgacgcgc atccgagatc gcaactggta
     1201 cgttcagccg tcttgcgcta caaccggcga cggactctac gaagggctca cctggctcac
     1261 gtccaactac aaatcctaac aactttatta tacatagttg ataattcact ggccgtcgtt
     1321 ttacggtacc atcgatgatg atccagacat gataagatac attgatgagt ttggacaaac
     1381 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
     1441 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
     1501 gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg
     1561 tggtatggct gattatgatc ctctagatca gatctgcgaa gatacggcca cgggtgctct
     1621 tgatcctgtg gctgattttg gactgtgctg ctcgcagctg ctgatgaatc acatacttcc
     1681 tccattttct tccactgatt gactgttata atttccctaa tttccaggtc aaggtgctgt
     1741 gcattgtggt aatagatgtg acatgacgtc acttccaaag gaccaatgaa catgtctgac
     1801 caatttcata taatgtgaaa acgattttca taggcagaat aaataacatt taaattaaac
     1861 tgggcatcag cgcaattcaa ttggtttggt aatagcaagg gaaaatagaa tgaagtgatc
     1921 tccaaaaaat aagtactttt tgactgtaaa taaaattgta aggagtaaaa agtacttttt
     1981 tttctaaaaa aatgtaatta agtaaaagta aaagtattga tttttaattg tactcaagta
     2041 aagtaaaaat ccccaaaaat aatacttaag tacagtaatc aagtaaaatt actcaagtac
     2101 tttacacctc tggttcttga ccccctacct tcagcaagcc cagcagatcc actagttcta
     2161 gagcggccgc caccgcggtg gagctccagc ttttgttccc tttagtgagg gttaattgcg
     2221 cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     2281 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     2341 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     2401 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     2461 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     2521 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     2581 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     2641 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     2701 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     2761 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     2821 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     2881 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     2941 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     3001 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     3061 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     3121 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     3181 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     3241 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg cattttggtc
     3301 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     3361 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     3421 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     3481 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     3541 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     3601 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     3661 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     3721 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     3781 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     3841 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     3901 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     3961 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     4021 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     4081 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     4141 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     4201 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     4261 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     4321 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     4381 gtgccaccta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     4441 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     4501 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     4561 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca cgctggctaa
     4621 gaactcatca gcctccccgg tccatctacc cacgtaccaa tgcaccaatt ggccacaatg
     4681 acggctacta catggtgcca ttccttcctc tttataggaa tggagactac ctcctgtcca
     4741 acaatgctct tggatacgag tacgcctacc tgttggaccc aggtcattgc acaacaccag
     4801 aaatgccctc tgatctgcaa aagacgtgaa tatctgttca gacacccata tccactctgt
     4861 tccacacagg tcagaggttt gtccaggagt tcttgacaga ggtgtaaaaa gtactcaaaa
     4921 attttactca agtgaaagta caagtactta gggaaaattt tactcaatta aaagtaaaag
     4981 tatctggcta gaatcttact tgagtaaaag taaaaaagta ctccattaaa attgtacttg
     5041 agtattaagg aagtaaaagt aaaagcaaga aagaaaacta gagattcttg tttaagcttt
     5101 taatctcaaa aaacattaaa tgaaatgcat acaaggtttt atcctgcttt agaactgttt
     5161 gtatttaatt atcaaactat aagacagaca atctaatgcc agtacacgct actcaaagtt
     5221 gtaaaacctc agatttaact tcagtagaag ctgattctca aaattgttag tgtcaagcct
     5281 agctcttttg gggctgaaaa gcaatcctgc agtgctgaaa agcctctcac aggcagccga
     5341 tgcgggaaga ggtgtattag tcttgataga gaggctgcaa atagcaggaa acgtgagcag
     5401 agactccctg gtgtctgaaa cacaggccag atgggccctc gagcaggaaa cagctatgac
     5461 catgattacg ccaagctatc aactttgtat agaaaagttg gcacgcgcag attccgtgtg
     5521 ggcctggtaa tatcttacca ttttaagcta ttttaagatt agggaaaaaa attaaataaa
     5581 actactttaa aattactttc cttaaaaagc aacatattta tttagttttt gtttaactgg
     5641 ctcaagattg taatatttta cttttacatg tttttttaag tacctaaatg tgttcaaaag
     5701 taaagtatgt tattgttttt tttttttaat gttagctaag ctttaaattg taatatatta
     5761 cttttaaaag tgacttttta aaaatgtaaa taaaaaagta actataaatg tgtaatgcat
     5821 gttatttctg tgggttttta attttcatct caactgggct tacaatttta acataataaa
     5881 aatcaaatca aatcaaatca ggcccttatt tgtcacatac actcttgtat actgaaataa
     5941 actaaatgtt gagaaaaatt taaatacacc aaaataacgc ataccttttt tgtaattacc
     6001 taagtaaaac atttagttcc ttttttaagt gactcaagat tgtaatctat taattgtaaa
     6061 agataaaact agtgatcaat ctgacgtgct tctatgcagc ctgtgagatt aattgtgatg
     6121 tttaattatt aaacaattta aatggatcaa acatttcaca tgtgcttatt gtgtccaaca
     6181 aaaccagcag aagcacttaa aaatattgca tgacaacatg agtttttaat gtaaattata
     6241 tatacttcct aaacgaagtt aatcatattt ctaaaataat aatgtcattg aaatatttaa
     6301 ataaatgata agctttagtt gattcaacat aaaattgaaa gcaataagga cttttttttt
     6361 ttacagcggt gaagcaaaag catgtaataa tcatataata taataaataa ataaataaat
     6421 aatggctctt ggtgcaaata tgaacacttt aaataaatat agaataaata tgatcaataa
     6481 gtcataacga tgtgttttat gttgattata atttaaacat gtttctaacg caagttttaa
     6541 aaactgtttt aaagtattat tttaagttat aagatcaatt tgactcaaca ttttgtagtg
     6601 aaaatatgtt gtgtaaaatt aatataatat acactataaa aatgcagggt tccacacaat
     6661 acattaatgt tgtcccaaca caaatcgatt aagttaacaa atatatgtgg attgaacata
     6721 aaaaattaag ttgtcccaat gaaatctcaa gaattgtgtt gtttcagctc attttaaata
     6781 agtagtttta acaagtaaaa aataacatat ttttgagtgt gtatatatat atattacaaa
     6841 atataataat ataaaatgta aaagatataa accatgctac atatatttaa gagctatttt
     6901 gcacattact tttttgcaag ccattttagt gtttttttta caaatatata ttagctaaat
     6961 atacaatatc ttaaatatac aattaatcag acatgcaatt aatgcattta acctacataa
     7021 aaatacatga tcaacatatg ttttatgctg attacaacta aatcaaacgt aatgaaatta
     7081 atttataaaa gcgatttaaa tatgtcaata ttttaagtta aaatcacttg aaacctacta
     7141 taaatgaact taaattaaaa aaatctcctt taattccaat gcaacagtgt tgttttttca
     7201 gctgaggacc cccggtgcag atggttggcc cccggcccgc tattctaagg aagcctttct
     7261 taattttagg gcggcgctgc atgcgctcag atcagattca cgggtggaga gggttatttt
     7321 ctctttttta ttttttcttg cctgctcata tttgggcctg tgtgaacccc aggctccgcc
     7381 tcggccccca ggagccgctc ttatttaaag gggatcctcg ccttcagtct tcgtcccagt
     7441 ggaagcatca gtcttgtgat aacaacttgg tgagtcctac agaccagcgc tcgactctag
     7501 gctttaaatg cattgatttc tctggattag ctaccgggag agttttaact ttgcttatta
     7561 gtcttgtgct cgaactgcta ggtttaaaga gtgttttctc taatgcatgc atgaagcgac
     7621 agctgaaaac cagaaataga gcgtttgtcc tcctgttaaa ctctagcctt tctttggcat
     7681 ttgtcagtcc gagtcttcag ctgccggcag gagattgggt ttcagtaaca cttgctgagc
     7741 acaaactaca cgtgctttct tcagatttaa ttcattatta atcattctag atttaaatga
     7801 tgttcttact cattgctgat tattttagcc ttgcggagca gattaaatga gcacgtgttt
     7861 cattaagatt gatttaaata taattaatat ttcaccactg tagttttaaa attgccttct
     7921 aagtcatggt tgatttttaa taggtatcgg ctctaattag attactattt tggtttttaa
     7981 atgttatttc tatttcattt gtaggctcgt ttttaatgtg attttattag attctagttt
     8041 taaaatcatc tcttagcctt aatgagcaga ctgaatgaac atgtttcatt aagattaact
     8101 tcattataat gaattattta atcattctag tgggttataa atcagctcta tttaaattaa
     8161 ttgtcgtgtt tatttctatt taatttttga ttattttttc agggtcgttt ttatgtgatt
     8221 ttttcccgtt tctaatttta aactaattgt ttgattttta tatgattcga ctctttttaa
     8281 atgaattgtt ttgctttttt tattgagcat atatatatta tacaatttca atcaacaatt
     8341 gttgtgcttt ttaaatgttg tttctatttc atttctaggg tcgattttat tcgattctag
     8401 ttttaaaatc gtgattttta acttctaacc ttactgagca gactgaatga acatgtttca
     8461 tcaagattaa cttcattata gttaattatt gaatcgtgct agtggtttat aaatcagctc
     8521 tatttaaatt aatcatcatc attatttcta tttaattttt attatttctt taggattatt
     8581 tttatgtgat ttttttcacg ttctagtttt aaactcaatg ctcattactg ctcacccttg
     8641 ctgagcagtc tttcgttaag attaaattaa ttaataacca ttctagtttt aaaacagtgt
     8701 cttaaggcat agctgatatt ttacaagttt actatttaat atatctcctt tcttttttta
     8761 gggtcgattt catctgattt gattaatttt tattatcttc actaagcttt ttgttctagg
     8821 ttttattctc agtattatta ctgttatatt tgaactgaat ttctcctcat agtttattca
     8881 ttttatttat ttattttttg tttattcgct attttgtatt ttagttttta attagaaatt
     8941 ttattagaat cgttttgaac tagtttttat gcactctaac attaatatta aatcataact
     9001 attattttta ttataattta tttgtgttta tctttttgtg tagctttcat gtgtttttat
     9061 tttagtttta ttatgtttgc attgaatttt ttggactagt ttatataacg atatacagaa
     9121 aagccatgac aacaaatgtt tttataatac ctattaagta taagctaatc agatttccac
     9181 tattctgttc aaagttaaca ttattagcca ttctgatgga tcatttgaca tttgtcatat
     9241 gtctttatga acactgctct ttcagatgcc acttttaaag ctgtttttta cagtaaatga
     9301 acgtttatta ttatgacatt atttcctaag gcctctagag tacagcagga tgtggcgtgt
     9361 gcattaccgt gtaattatgc tgactgtaaa tgtgcattta tagactgtca gcatgtctga
     9421 ctgtgcgctg ttgtgattgg tcctgcacag accaag
//
