LOCUS       sequence-384398-        8767 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8767)
  AUTHORS   Kemmler CL, Moran HR, Murray BF, Scoresby A, Klem JR, Eckert RL, 
            Lepovsky E, Bertho S, Nieuwenhuize S, Burger S, D'Agati G, Betz C, 
            Puller AC, Felker A, Ditrychova K, Botschi S, Affolter M, Rohner N, 
            Lovely CB, Kwan KM, Burger A, Mosimann C
  TITLE     Next-generation plasmids for transgenesis in zebrafish and beyond.
  JOURNAL   Development. 2023 Mar 28:dev.201531. doi: 10.1242/dev.201531.
  PUBMED    36975217
REFERENCE   2  (bases 1 to 8767)
  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: 
            "Development. 2023 Mar 28:dev.201531. doi: 10.1242/dev.201531."
COMMENT     Sequence Label: pCK004 ubi:mCerulean_SV40 (#302)
FEATURES             Location/Qualifiers
     source          1..8767
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    37..57
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    3592..3616
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      3639..3648
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3645..3647,3648..3650,3651..4163)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIE"
     primer_bind     complement(3690..3711)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3951..3970)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4298..4319
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(4381..4405)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    4407..4541
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(4604..4624)
                     /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(4632..4648)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    4664..4798
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(5237..5424,5425..5436)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5470..5486)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5470..5486)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(5523..5543)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5523..5541)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5562..5578)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5575..5597)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5586..5602
                     /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(5610..5616,5617..5634,5635..5640))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5655..5676
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5793..5810)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5964..6552)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6044..6063)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6723..7514,7515..7583))
                     /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     7346..7365
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7584..7688)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7851..7872)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(8221..8232,8233..8395)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
ORIGIN
        1 caggaaacag ctatgaccat gattacgcca agctatcaac tttgtataga aaagttggct
       61 ccgaattcgc ccttaaaact cgagaccagc aaagttctag aatttgtcga aacatttatg
      121 ttatatattt cctgaaaaaa attctgagta agttcttaag tgttattgcc agcaacataa
      181 acaacagacg gcaaaatgaa taaatgataa caaagcagta ggcttaaata aacctaattt
      241 ttataggctg ttctctacaa ccctcaaaca gtgattagtt ttgtacttat aaacttgccc
      301 tttcattcat atttcaagaa aattggttca gaagatctgg atattctagc agttgttcaa
      361 gctcatggag ggatcagtga cctgattcca caatgactag gcctaatcca gaaattagat
      421 gactgtcaac ataaaaaggc acagcactca ctagctgccc tatatatttt attatatttt
      481 acatatatta ttttatttat ttagctctga gtgctgtact ttctggttaa agaaaactgc
      541 ttacaacagc taacctgtac tacctcaggc tcagggaatt tggaacaggt ttgtctggtt
      601 tgtttcttta accatgcatg cttgttttca actatggcaa cacagtcaca tgggacatta
      661 cagaaatgat ttgtcgatga catgcgactt ttctttaata aagcgcaaag atcccaaaaa
      721 gcaaactttt aacaaaaatc atataattat attttcaatc cagctttgta gcaactttgt
      781 gctgctgttc actcagcaac agatagtcag tataaggtca gtgtgtctca aagcagtgcc
      841 atctgtttca cacattgcgt tctatatata agtgtgctgg ttgacacgac actgtataag
      901 gcctaggcta aaacacaaac aatgtagaat gacactgtgt tttttttgta aacaaatgtt
      961 gtttttggtt aaacatcttt gtgaaaacat cctcctgtca tgtatttgct atattcaaat
     1021 gttaaacccg tgcagaatag aacatataca aaaaaaaaca acacaacaca tttttaaaca
     1081 ttattaaata tcaagtattg ctggcagttc tgtttctgtt ttacagtacc ctttgccaca
     1141 gttctccgct tttcctggtc cagattccac aagtctgatt caccaatagc aaagcgaata
     1201 aacaaccaaa gcagccaatc actgcttgta gactgtcctg cgagaccggc ccattccagc
     1261 acattctgga aacttccttt atatgataat tataaataca tttaaattat tgatacaaaa
     1321 catgtaattc ctagaacata accatagcaa tcattagttt tcagggtaat tatgtatttt
     1381 taggatttga ctgcggaaag atctggtcat gtgacgtctc atgaacgtca cggccctggg
     1441 tttctataaa tacagtagga ctctcgacca tcggcagatt tttcgaagaa gaagatcagt
     1501 ttcaggagcc gtactgttcc gtttcaacgc aaatattaac ggtaagagcg aatttcctag
     1561 tttgttttca tgccattctt taaaaccata gcgtattact ttaattatag taaactttcg
     1621 ctttctttat tacaagagac gttttgtgtt gattctccgc ggacattttc ggtcagacaa
     1681 tcagaaaatg accgcggagg accagtaact tgcattacac gtaagttaaa tcttcgtgta
     1741 ttaaaatggt taggttgtta acgtcaaata ggttaccgtg tttgcgtgtg atcaggttgg
     1801 ttttgttaga tttttgtcag tatttttaat ttatttgttt tagtttattt attttttttg
     1861 ctgaatcata gtttgtgaac aaagaacccg gatgttacat acagtacagc cgccatgtta
     1921 cagagagtta taacttaatc attttaaaaa taattttgcc ttacttttag tttgtcatgt
     1981 tgagaaatga ggaaatgtta aaatgaggaa atatccaatt aatttaatat atcaaaataa
     2041 tccatgatta caatgcactg aactggagaa aattaagatg ttttctagtg tcatgaaaca
     2101 aatgtaagag atgtacattg tagatgtttt atgtcaagaa ttggctagtt gatgcagcat
     2161 actggcgata ctcagttgta ataacagtaa cgttacatgt taatagacta ctgagtatgc
     2221 tgttctgtct atgtatgctc tgtaagctac gagaaggact tttttaaaca gtaaagggtg
     2281 caatattttt acaaattgaa ttaaataaag gctgtctatt aagtaatatg cttgatattt
     2341 ttcttacttg atcggaaata agaaaaaata taaacgttgt tgctctaaaa atcctagttc
     2401 agtttagcca accacaaata cctttttgtt cctccaacag tttttttttc ttctctataa
     2461 tatttggcag tctatagtac tccaaatgtt tccccacagt ctaactaatt ggtacagcca
     2521 aaatcatgac acttattgca ataataattt tggttcattg gcattgttga tagcctgtgc
     2581 cactaatatg gtcgattgat catgcttcag gaagaaaact atattgtttg atgtaagatt
     2641 attaaatctt cacctgcttc cattacaaac tattcccatc ttattgaatt ctggtatgtc
     2701 ttaaaggatt agttcacttc ccaaatcaaa atttacttag ccttttttca tccatgatcc
     2761 ctttttttca tcattaatga agaaattgtt tttgaaaaag tttcaagatt tttttctcta
     2821 tattgtggag cttgttagtt taaaattcca aaatgcaata tgtggcttca aatggttcta
     2881 aatgatccca gtcaaggaat aacagtctta tctaatgaaa ccattagacc tttaaaaata
     2941 aaaaaaataa aagtatttat ttttaaatga ctgagtgatt aagttgaatt tcagcgtttc
     3001 cttactgtgt agaagtcctt ccttactggc cccacccttt ggttctctgc caatctgcta
     3061 cctaatgtaa tgttgtggaa cattattatt ctttatttct taatttttta ttttttattt
     3121 taaaacaatg taaactgcac agatgtgcag tttgtttaaa atggccaatg ctttggaaat
     3181 gcatgacata attagatttc atgatgcaca aagccaaatc tcagagcttg tgcaaaatga
     3241 gctatcattt cactaggtaa gaccctaaat tttcatatag gatcatttgg acaattttgc
     3301 tgcaggtaaa atgcattcta tagtccactg tcagccattg ttttggatag tatttatttt
     3361 tctctacaag tatagtcaat agttttctat tattttaaag gtttgtaaca tttaagggtg
     3421 accaaatgca aagtaaaatt tcattttcgg gtgaactatc tcgtttaaca tgggagaagt
     3481 gcaaaacata cattattggc tagaacattg tagtattttt taaatggaaa tgtgtgattg
     3541 ctaatcttac tttgaatttg tttacaggga tcctttaagg gcgaattcga cccaagtttg
     3601 tacaaaaaag caggctccgc ggccgccccc ttagatccgc caccatggtg agcaagggcg
     3661 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     3721 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     3781 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga
     3841 cctggggcgt gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca
     3901 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     3961 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     4021 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaacg
     4081 ccatcagcga caacgtctat atcaccgccg acaagcagaa gaacggcatc aaggccaact
     4141 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     4201 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     4261 ccaagctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     4321 ccgccgccgg gatcactctc ggcatggacg agctggccct cgagaagggt gggcgcgccg
     4381 acccagcttt cttgtacaaa gtgggggatc cagacatgat aagatacatt gatgagtttg
     4441 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     4501 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     4561 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttccaacttt attatacata
     4621 gttgataatt cactggccgt cgttttacgg taccatcgat gatgatccag acatgataag
     4681 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     4741 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     4801 caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta
     4861 aagcaagtaa aacctctaca aatgtggtat ggctgattat gatcctctag atcagatctg
     4921 cgaagatacg gccacgggtg ctcttgatcc tgtggctgat tttggactgt gctgctcgca
     4981 gctgctgatg aatcacatac ttcctccatt ttcttccact gattgactgt tataatttcc
     5041 ctaatttcca ggtcaaggtg ctgtgcattg tggtaataga tgtgacatga cgtcacttcc
     5101 aaaggaccaa tgaacatgtc tgaccaattt catataatgt gaaaacgatt ttcataggca
     5161 gaataaataa catttaaatt aaactgggca tcagcgcaat tcaattggtt tggtaatagc
     5221 aagggaaaat agaatgaagt gatctccaaa aaataagtac tttttgactg taaataaaat
     5281 tgtaaggagt aaaaagtact tttttttcta aaaaaatgta attaagtaaa agtaaaagta
     5341 ttgattttta attgtactca agtaaagtaa aaatccccaa aaataatact taagtacagt
     5401 aatcaagtaa aattactcaa gtactttaca cctctggttc ttgaccccct accttcagca
     5461 agcccagcag atccactagt tctagagcgg ccgccaccgc ggtggagctc cagcttttgt
     5521 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
     5581 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     5641 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     5701 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5761 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5821 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     5881 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5941 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     6001 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     6061 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     6121 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     6181 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6241 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6301 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6361 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     6421 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6481 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6541 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6601 aactcacgtt aaggcatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6661 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6721 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6781 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6841 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     6901 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     6961 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     7021 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     7081 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     7141 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7201 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7261 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7321 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7381 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7441 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7501 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7561 acacggaaat gttgaatact catactcttc ttttttcaat attattgaag catttatcag
     7621 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7681 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     7741 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     7801 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     7861 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     7921 agggcgatgg cccacgctgg ctaagaactc atcagcctcc ccggtccatc tacccacgta
     7981 ccaatgcacc aattggccac aatgacggct actacatggt gccattcctt cctctttata
     8041 ggaatggaga ctacctcctg tccaacaatg ctcttggata cgagtacgcc tacctgttgg
     8101 acccaggtca ttgcacaaca ccagaaatgc cctctgatct gcaaaagacg tgaatatctg
     8161 ttcagacacc catatccact ctgttccaca caggtcagag gtttgtccag gagttcttga
     8221 cagaggtgta aaaagtactc aaaaatttta ctcaagtgaa agtacaagta cttagggaaa
     8281 attttactca attaaaagta aaagtatctg gctagaatct tacttgagta aaagtaaaaa
     8341 agtactccat taaaattgta cttgagtatt aaggaagtaa aagtaaaagc aagaaagaaa
     8401 actagagatt cttgtttaag cttttaatct caaaaaacat taaatgaaat gcatacaagg
     8461 ttttatcctg ctttagaact gtttgtattt aattatcaaa ctataagaca gacaatctaa
     8521 tgccagtaca cgctactcaa agttgtaaaa cctcagattt aacttcagta gaagctgatt
     8581 ctcaaaattg ttagtgtcaa gcctagctct tttggggctg aaaagcaatc ctgcagtgct
     8641 gaaaagcctc tcacaggcag ccgatgcggg aagaggtgta ttagtcttga tagagaggct
     8701 gcaaatagca ggaaacgtga gcagagactc cctggtgtct gaaacacagg ccagatgggc
     8761 cctcgag
//
