LOCUS       sequence-331454-        5568 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express EGFP-zebrafish Plaat1 (WT) in zebrafish lens.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5568)
  AUTHORS   Morishita H, Eguchi T, Tsukamoto S, Sakamaki Y, Takahashi S, Saito 
            C, Koyama-Honda I, Mizushima N
  TITLE     Organelle degradation in the lens by PLAAT phospholipases.
  JOURNAL   Nature. 2021 Apr;592(7855):634-638. doi: 10.1038/s41586-021-03439-w.
            Epub 2021 Apr 14.
  PUBMED    33854238
REFERENCE   2  (bases 1 to 5568)
  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"; doi: 
            "10.1038/s41586-021-03439-w"; journalName: "Nature"; date: 
            "2021-04"; volume: "592"; issue: "7855"; pages: "634-638"
COMMENT     Sequence Label: Tol2-aACry::EGFP-zebrafish Plaat1 (WT)
FEATURES             Location/Qualifiers
     source          1..5568
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1010..1030
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1012..1030
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     1067..1083
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1067..1083
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     misc_feature    join(1117..1128,1129..1316)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     promoter        1902..1920
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1902..1919
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      1979..1988
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1985..1987,1988..1990,1991..2701)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1985..1987,1988..1990,1991..2701)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2030..2051)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2291..2310)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2638..2659
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3477..3611
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3501..3520)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3555..3574
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(3716..3853,3854..3865)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     promoter        complement(3896..3914)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3924..3941)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3924..3940)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3933..3955)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4249..4270
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4433..4537
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4538..4606,4607..5398)
                     /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(4756..4775)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
     1021 tcactaaagg gaacaaaagc tggagctcca ccgcggtggc ggccgctcta gaactagtgg
     1081 atctgctggg cttgctgaag gtagggggtc aagaaccaga ggtgtaaagt acttgagtaa
     1141 ttttacttga ttactgtact taagtattat ttttggggat ttttacttta cttgagtaca
     1201 attaaaaatc aatactttta cttttactta attacatttt tttagaaaaa aaagtacttt
     1261 ttactcctta caattttatt tacagtcaaa aagtacttat tttttggaga tcacttgggc
     1321 ccggtcgaca ttaatagtgt gcattcagtg caggatgtta tgtatttatt ttctgtagtg
     1381 caaacttttt caaagaattt tccagtctaa gcttttcagt caagaaaaaa aaaagcgatt
     1441 gattatctgt gatatttaaa aaaattagct cttaaatctg gtgaaccaag tggccacaac
     1501 acattaaata ctgacgttct acagccatgt agcttcatct ggtctggttt gttttggcag
     1561 gcacttattt tccggtgata gttggccaca agagtattgt ctgagccatt cagtgctaga
     1621 ctgtcattct caggtcagag tccaccccgc tgattctgct gacaacactg ttcccaccat
     1681 gagataatgc cattccagag agatccattt gtaagcccct ctttctgcag cacaggtata
     1741 taaccagggg tctgcctcca ctaaggccgg cacacatcat ttggggatct ttgtactgta
     1801 ctagcctcta ttagcctcct acttgcgttc agttcacatc ttttgttcat caggtgcaat
     1861 aagttacagt aaggagttac caggtctgac agatctgctt gatttaggtg acactataga
     1921 atacaagcta cttgttcttt ttgcaggatc tcgagaccat ggaatcgata taaagcttgc
     1981 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     2041 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     2101 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     2161 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
     2221 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     2281 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     2341 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     2401 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     2461 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     2521 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     2581 ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat
     2641 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     2701 gggttcaggc tcgggatcca tggataatca acagagaaga actgataata tggcgtcagg
     2761 tgaaactgat caccttcagt gtgtggagga gcctcagcca ggagacctca tcgagatctt
     2821 cagaccggcc tatcagcact gggctctgta tctgggagac ggatacatca tcaacctcac
     2881 gcctgtcgat gaaggtcagg ccactgctgt gtccagtgtg aagtcggtgt tcagccgtaa
     2941 agctgtggtc cgcatgcagc tcttgaagga ggtggtgggt gcagattcat acaggatcaa
     3001 taataaatat gatgacgatc acacgccgct gcccgtctcg gagatcatcc agcgggcaca
     3061 gatgctcatc ggccaggagg tgtcgtatga cctgctgggc agcaactgtg agcacttcgt
     3121 cactctgctc cgctacggtg aaggagtgtc cgaacaggca tcgcgggcta tcggtgccat
     3181 cagtctggtg acagcagccg caagtgcttt ctcagtgctg ggtctcatca acacccgatc
     3241 cagaaacaga cccttctagc tcgagagcgg ccgctacgta aattccgccc ccccccctaa
     3301 cgttactggc cgaagccgct tggaataagg ccggtgtgcg tttgtctata tgttattttc
     3361 caccatattg ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg tcttcttgac
     3421 gagcattcct aggggatccc tcgagcctct agaactatag tgagtcgtat tacgtagatc
     3481 cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
     3541 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     3601 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt
     3661 gggaggtttt ttaattcgcg gccgcggcgc caatgcattg ggcccggtaa gatctaatac
     3721 tcaagtacaa ttttaatgga gtactttttt acttttactc aagtaagatt ctagccagat
     3781 acttttactt ttaattgagt aaaattttcc ctaagtactt gtactttcac ttgagtaaaa
     3841 tttttgagta ctttttacac ctctgtcaag aaccatatgc cggtacccaa ttcgccctat
     3901 agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     3961 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     4021 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     4081 gacgcgccct gtaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     4141 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     4201 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     4261 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     4321 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     4381 ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac
     4441 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     4501 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     4561 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     4621 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     4681 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     4741 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     4801 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     4861 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     4921 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     4981 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     5041 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     5101 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     5161 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     5221 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     5281 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     5341 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     5401 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     5461 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     5521 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttc
//
