LOCUS       sequence-373955-        5994 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Intron-based gene-break transposon (GBT) for an effective and 
            revertible loss-of-function tool for zebrafish. Lens tagBFP 
            frameshift 3..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5994)
  AUTHORS   Ichino N, Serres MR, Urban RM, Urban MD, Treichel AJ, Schaefbauer 
            KJ, Greif LE, Varshney GK, Skuster KJ, McNulty MS, Daby CL, Wang Y, 
            Liao HK, El-Rass S, Ding Y, Liu W, Anderson JL, Wishman MD, 
            Sabharwal A, Schimmenti LA, Sivasubbu S, Balciunas D, Hammerschmidt 
            M, Farber SA, Wen XY, Xu X, McGrail M, Essner JJ, Burgess SM, Clark 
            KJ, Ekker SC
  TITLE     Building the vertebrate codex using the gene breaking protein trap 
            library.
  JOURNAL   Elife. 2020 Aug 11;9. pii: 54572. doi: 10.7554/eLife.54572.
  PUBMED    32779569
REFERENCE   2  (bases 1 to 5994)
  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: "2020-08-11"; volume: "9. pii: 54572. doi"; pages: " 
            10.7554/eLife.54572"
COMMENT     Sequence Label: pGBT-RP8.3
FEATURES             Location/Qualifiers
     source          1..5994
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    join(10..21,22..184)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     promoter        complement(204..222)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     protein_bind    complement(248..281)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             451..1125
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="ASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAKL
                     KVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKMR
                     LKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHST
                     GA"
     CDS             451..1125
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="ASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAKL
                     KVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKMR
                     LKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHST
                     GA"
     primer_bind     complement(578..596)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     846..865
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1040..1063
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             3000..3020
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3024..3725
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     protein_bind    complement(3891..3924)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     3958..3977
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3958..3976
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(3999..4186,4187..4198)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(4252..4271)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4263..5072
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4330..4349)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      5215..5803
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5704..5723
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5957..5974
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ggtaccgagc agaggtgtaa aaagtactca aaaattttac tcaagtgaaa gtacaagtac
       61 ttagggaaaa ttttactcaa ttaaaagtaa aagtatctgg ctagaatctt acttgagtaa
      121 aagtaaaaaa gtactccatt aaaattgtac ttgagtatta aggaagtaaa agtaaaagca
      181 agaaagaaaa ctagagtggt ctccctttag tgagggttaa ttgatgggat agggataaca
      241 gggtaatata acttcgtata gcatacatta tacgaagtta tcgttaccac ccactagcgg
      301 tcagactgca gattgcagca cgaaacagga agctgactcc acatggtcac atgctcactg
      361 aagtgttgac ttccctgaca gctgtgcact ttctaaaccg gttttctcat tcatttacag
      421 cagccggaat tcacccgggt accggaaggt gcctcctccg aggacgtcat caaggagttc
      481 atgcgcttca aggtgcgcat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
      541 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggc
      601 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tccagtacgg ctccaaggcc
      661 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc
      721 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc
      781 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
      841 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctccaccga gcggatgtac
      901 cccgaggacg gcgccctgaa gggcgagatc aagatgaggc tgaagctgaa ggacggcggc
      961 cactacgacg ccgaggtcaa gaccacctac atggccaaga agcccgtgca gctgcccggc
     1021 gcctacaaga ccgacatcaa gctggacatc acctcccaca acgaggacta caccatcgtg
     1081 gaacagtacg agcgcgccga gggccgccac tccaccggag cctaagacta gtaattaagt
     1141 ctcagccacc gttaactgaa catgtcaaaa cctgtggaga ctgttgagat ttgatgttct
     1201 gaaaagataa agcctataaa taaaatgttg cccaaatttc ctgcctgatg tttttctttg
     1261 tctttgctac atggctttgc tgctcggatc ggctcactct gtgtatgcca cgttcacttt
     1321 gtactctcct tctcacggta ggtttattat ttttagatgt gcagttagtt tctgtgaaat
     1381 aacacaccac acactgatat tgtctgtgca ttgacttggt gagtgcacat tgtttttgat
     1441 cttgacatat ttatatttga ttgatcaggt gaactgtgtg aatctaaagt gctccataca
     1501 gatgttctgc attgaaaata ttctcatttt attagtggaa gtgagtgtat gccacatcca
     1561 atcaatttca gcaaacaccc cagtatgatt taatgcaaaa aaatgaaggt atcaaacacg
     1621 cattactact ttgcagttaa atatttaaca tttattccaa cacgaaaaaa agcagtaaat
     1681 aacactttga caaacacgtc aggacatctt atttttgtca ccctcacagg caatttagta
     1741 taatatatta tatatatata tatatcatat aataatattc agtataatat atatatatat
     1801 atcatattat aatattcagt ataatataaa acacaaacac atatatgtat aatataatat
     1861 aacattttta tttattgaga tgcctctatg gaccgtgtta taagaagtaa agatcaggag
     1921 aagtaaacat gaagtgtaat tatgaatact gatgttaaat taagctatga tgagttttca
     1981 ctgttaattt accatctcaa ttaaatgttg atgcctccat gaccaagtta agcagatgag
     2041 actgaggaca actgtagaag acaagatgtt cactttgctg aatatagctg gcttgacagt
     2101 tatctatgac tctataaata tatatatatt ttttttttta taaaatgatt tatttataac
     2161 tatatatcca tttctcagac aggtgcttca tatccctcac tcccgtagct gtccatgctg
     2221 gatctgtccc cgttgttttt aaaaagctaa ataagttatt aacatgactg catccagcga
     2281 gccaaacctg tctggtgtac agctaccaga gaagcttgag atcctagtca ccggcgttcc
     2341 tagttcttta aactgttaca aagtgtttgg accagcaact gaattgcact gtaaattacc
     2401 acatcagggt ctccggggtc aacgctgcta agcaatggac tggaaaattt gtgacattga
     2461 gtttttggaa taactgggaa tgctggataa ctctttgggt aaagctatgg aacaaaagta
     2521 ccagcatgaa ctaagaaatg actgtcacag cacaacatca aacacaatgc aagacaaagg
     2581 ttctctgctt ttgccttact tgacagaaaa agtaagctct aggttcatcg catgcagaca
     2641 gcagcaatac atgacatata taaaggctgg ctcttttctc ccttccattg aactgaaact
     2701 tccactcagt cagacttgcg ggggatccac agcctgacta actcgacaaa cccccccaaa
     2761 cctaaggtga gttgatcttt aagcttttta cattttcagc tcgcatatat caattcgcgc
     2821 gcattgtcac actagcaccc actagcggtc agactgcaga ttgcagcacg aaacaggaag
     2881 ctgactccac atggtcacat gctcactgaa gtgttgactt ccctgacagc tgtgcacttt
     2941 ctaaaccggt tttctcattc atttacagtt cagccggaat tcacctcgag accatggctc
     3001 caaagaagaa gcgtaaggta accatgagcg agctgattaa ggagaacatg cacatgaagc
     3061 tgtacatgga gggcaccgtg gacaaccatc acttcaagtg cacatccgag ggcgaaggca
     3121 agccctacga gggcacccag accatgagaa tcaaggtggt cgagggcggc cctctcccct
     3181 tcgccttcga catcctggct actagcttcc tctacggcag caagaccttc atcaaccaca
     3241 cccagggcat ccccgacttc ttcaagcagt ccttccctga gggcttcaca tgggagagag
     3301 tcaccacata cgaagacggg ggcgtgctga ccgctaccca ggacaccagc ctccaggacg
     3361 gctgcctcat ctacaacgtc aagatcagag gggtgaactt cacatccaac ggccctgtga
     3421 tgcagaagaa aacactcggc tgggaggcct tcaccgagac gctgtacccc gctgacggcg
     3481 gcctggaagg cagaaacgac atggccctga agctcgtggg cgggagccat ctgatcgcaa
     3541 acatcaagac cacatataga tccaagaaac ccgctaagaa cctcaagatg cctggcgtct
     3601 actatgtgga ctacagactg gaaagaatca aggaggccaa caacgagacc tacgtcgagc
     3661 agcacgaggt ggcagtggcc agatactgcg acctccctag caaactgggg cacaagctca
     3721 attaaagctg agatctacct aaggtgagtt gatctttaag ctttttacat tttcagctcg
     3781 catatatcaa ttcgaacgtt taattagaat gtttaaataa agctagatta aatgattagg
     3841 ctcagttacc ggtctttttt ttctcattta cactgagctc aagacgtctg ataacttcgt
     3901 atagcataca ttatacgaag ttattaccct gttatcccta tggctagcga tgggatctaa
     3961 tacgactcac tatagggaga ccaagggaaa atagaatgaa gtgatctcca aaaaataagt
     4021 actttttgac tgtaaataaa attgtaagga gtaaaaagta cttttttttc taaaaaaatg
     4081 taattaagta aaagtaaaag tattgatttt taattgtact caagtaaagt aaaaatcccc
     4141 aaaaataata cttaagtaca gtaatcaagt aaaattactc aagtacttta cacctctggg
     4201 atccagctgc atcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg
     4261 ttatgagcca tattcaacgg gaaacgtcga ggccgcgatt aaattccaac atggatgctg
     4321 atttatatgg gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc
     4381 gcttgtatgg gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg
     4441 ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc
     4501 cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc
     4561 ccggaaaaac agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg
     4621 atgcgctggc agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta
     4681 acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg
     4741 atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa
     4801 tgcataaact tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg
     4861 ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa
     4921 tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt
     4981 cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc
     5041 agtttcattt gatgctcgat gagtttttct aatcagaatt ggttaattgg ttgtaacact
     5101 ggcaaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     5161 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     5221 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     5281 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     5341 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     5401 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     5461 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     5521 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     5581 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acggttcccg aagggagaaa
     5641 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     5701 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     5761 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     5821 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     5881 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     5941 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaacagattt aaat
//
