LOCUS       sequence-182768-        7941 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  overexpression of microRNA and Dendra2 in Zebrafish.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7941)
  AUTHORS   Hsu AY, Wang D, Liu S, Lu J, Syahirah R, Bennin DA, Huttenlocher A, 
            Umulis DM, Wan J, Deng Q
  TITLE     Phenotypical microRNA screen reveals a noncanonical role of CDK2 in 
            regulating neutrophil migration.
  JOURNAL   Proc Natl Acad Sci U S A. 2019 Aug 26. pii: 1905221116. doi: 
            10.1073/pnas.1905221116.
  PUBMED    31451657
REFERENCE   2  (bases 1 to 7941)
  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: "Proc Natl 
            Acad Sci U S A. 2019 Aug 26. pii: 1905221116. doi: 
            10.1073/pnas.1905221116."
COMMENT     Sequence Label: Tol2-lyzC-microRNA 1388-Dendra2
FEATURES             Location/Qualifiers
     source          1..7941
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(119..140)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(536..547,548..685)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     regulatory      3113..3122
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4430..5122
                     /codon_start=1
                     /product="monomeric photoswitchable green-to-red 
                     fluorescent protein from Dendronephthya"
                     /label=Dendra2
                     /note="mammalian codon-optimized"
                     /translation="MNTPGINLIKEDMRVKVHMEGNVNGHAFVIEGEGKGKPYEGTQTA
                     NLTVKEGAPLPFSYDILTTAVHYGNRVFTKYPEDIPDYFKQSFPEGYSWERTMTFEDKG
                     ICTIRSDISLEGDCFFQNVRFKGTNFPPNGPVMQKKTLKWEPSTEKLHVRDGLLVGNIN
                     MALLLEGGGHYLCDFKTTYKAKKVVQLPDAHFVDHRIEILGNDSDYNKVKLYEHAVARY
                     SPLPSQVW"
     polyA_signal    5233..5354
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5270..5289)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5324..5343
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    join(5478..5665,5666..5677)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5732..5752)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5732..5750)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5771..5787)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5771..5787)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5784..5806)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5795..5811
                     /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(5819..5825,5826..5843,5844..5849))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5864..5885
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6002..6019)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6173..6761)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6253..6272)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6932..7723,7724..7792))
                     /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     7555..7574
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7793..7897)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
       61 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
      121 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
      181 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
      241 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
      301 gggaaagccg gccacaatga cggctactac atggtgccat tccttcctct ttataggaat
      361 ggagactacc tcctgtccaa caatgctctt ggatacgagt acgcctacct gttggaccca
      421 ggtcattgca caacaccaga aatgccctct gatctgcaaa agacgtgaat atctgttcag
      481 acacccatat ccactctgtt ccacacaggt cagaggtttg tccaggagtt cttgacagag
      541 gtgtaaaaag tactcaaaaa ttttactcaa gtgaaagtac aagtacttag ggaaaatttt
      601 actcaattaa aagtaaaagt atctggctag aatcttactt gagtaaaagt aaaaaagtac
      661 tccattaaaa ttgtacttga gtattaagga agtaaaaggc tagcaagatc tgctcgagat
      721 acatttgatt agaagagttt gacttgggtt gattatagca aatagtggaa cacaaacaga
      781 ttttgaaata ttcttgcaaa tttatttatt aaatccattc aaatattgtc attttttgta
      841 ctgtgtaatt tatattgatt tgctcataca atatttacat ttcgcttaaa caatacacta
      901 ttatttatat ttaaaaatag ctatatataa atcaaatgta tagctctttt ttacacaatt
      961 aatcaaagct aaaatatttg tcacacaaat acagtaagcc acttcacaat gatatcttca
     1021 aaatctgtgc ttgactgaat catttcattt attgttttgt aattcacatt ataccttacg
     1081 ttctccaggc acagtttgaa ttgttgataa atgaagataa tacatctgat ttaattagtc
     1141 tcctttgttt ataataactt taatctaatt tgatagtaat aattccagca atacatccat
     1201 tacaataaaa atcgcttttt tattttgcac attagtttgc cggtatttgt ttaaagtttc
     1261 accatacttt taaatcatgt gctatttgaa aagggctgtt atgactggaa ttaaatgttt
     1321 ttttttatat cagtgtttta gattgaagta caatatttat aggtaaaatt caccagcata
     1381 tgcttaaaga tagccatgct ccaggtcctt gtgtccttct aaggtctgtt tgtggtttaa
     1441 gctgtttgtt gacacacgct tgtcatctcc atttttctgt gaagttattg tcagtttatt
     1501 gcacactggg ttgcattaag ctgcagtttt ttcacaacag ttttccacat tgtaacagtc
     1561 agtctccaaa ttattccatc agtcctcaaa tttagtcatt tgaggccatg aaaacactta
     1621 aaactacatg aaggtagttt ggcattaggt tttagacttg ttcttgtttg gcatattgca
     1681 acctgtgtaa aaatgatttg atttttattt tttttttttt tttttcagaa agaaacggtc
     1741 caaaaagagt tatgtgaaca tttatccatg tgtttaatgt tttttaaatt aaaaaaaaac
     1801 atttttatta aaccacttgt gactaaacct tttttagtta ttctattttg tgacgagtct
     1861 gaatggccaa agtactcttg aatccaaaat ctttatagta aaatgtttaa tcttacttgc
     1921 agtgttgggt ataatttgta acaaagttga attccatatc tgctgagtga aaaagtaaag
     1981 taagagattg ctgtaaatat ttaggttatt tatttattaa tacaatatag aaagtaacaa
     2041 aacacaatgt aaaaaatcaa taaagaaagt aaatattaaa agtaatgata aactaatgat
     2101 gtaaacaata ttagcaacaa agtttgtaca attcagggag ggtaaagaat ttcacctaat
     2161 agtcaaactc tgtattaaat aatataaaat ctttgttctt agtctgaaga ggagactgtg
     2221 agagaaagct tactggttgt taaattagaa aatttgtaac ttgcagatta aaatgattta
     2281 aataatttta taaaggccat gtttgaaaca aatgtaaaaa cataataaat gttaagaatt
     2341 tcagatattg agattttaca atttaaaaaa agcaataaag taatagcaga gacctacact
     2401 cttaaagtaa attacccaac tctgcttact agtaactttt ggaatagtta ctagtatcca
     2461 ttgaacaaat actagaatct aattaaattg tgtgagtgtc caatgaaaag attcttgtat
     2521 ttaaaacagt accagtatct atatatgtac aaaatttagt gctagcaaaa tgcaaataaa
     2581 tactactgca tataatggat tgcattgtaa atcgactcgc cataatacat ttatttccta
     2641 tttttacttt taacaaaaaa taagttcact gagtatttca tgtcattaaa tcgcattaaa
     2701 cctcttgcat gctttatgta cactagtgca gttcgctaaa tgaccaccgg tgtcactaat
     2761 aacacgatcg tttctgaatt ctgcattaaa cgccatttat acgttgctct tattagtaaa
     2821 caatttctcc atcatgagct cgaccagtat tgcatatttc atcctcttaa atccagagtt
     2881 gtttctttta atcactttct cttcatgata aaagcagttt ttggttttca gaagtttaca
     2941 acaaacccag tcagttattg cctcagcaaa cactggagca actgcacact tgtgaaaagc
     3001 ttcttgcgaa acacaaacct ttctctggac ttataaataa aggcattcag cagctccacc
     3061 atacacctcc actggtcttg ataaagcaga tatcagcagt gatacaggta ccgccaccat
     3121 ggatgaggaa atcgctgccc tggtcgttga caacggctcc ggtatgtgca aagccggttt
     3181 tgctggagat gatgcccctc gtgctgtttt cccctccatt gttggacgac ccagacatca
     3241 ggtacattta ataaaccatc ctaataacat gttaatcata ttgaaccaca ttaaactctc
     3301 atttgtgcat tttacatctt ttaaaaccga tttatattcg tttttagata atgaagttta
     3361 ttaaagtgtt aattcacagc tttattgctt gttttgagaa tctgcttgca taagtgattt
     3421 ttaaacttga tctgattttc gtttgtttcg aagttcttca gctttgaaag tcagatcttc
     3481 aattcacagc catgccatag ttttaggata cagtataagt agacgaattg ggatgcatcg
     3541 tagattttat gctgtgtttg ttatggtttt tcagtggtag agattttcag agtattatta
     3601 gggttatagt cagtgtcagg tttatttatc gtcagcgtga tgggcaaaaa acagctggca
     3661 gtacgggctc aaacctgtct ttctttacct gttgatttta gatgcatttt cctgtattca
     3721 acaaaataat atttgaaaat aagcatttct ttcctctcat cgtttacttt gtgtgaaatt
     3781 gatactgtga gagtgtactt ttttttttat tgccattctt agaataattt tatttttttt
     3841 tttgactagg tgacgtacat cacataagta cttaatctga tggatgagca gtgctttcca
     3901 ggactgtcca acctgagaat gcttgagttt tggtcaatct caggttcgtc agcccatgaa
     3961 aaactgtctc gctcaaagcc acatggacga accgctgctc ctccaaacca cctaaacgcc
     4021 gcttaagacc cacatcaagc caactgctgt tactctcgcc cagaaatctg cgccaacatg
     4081 gacacaccca gtggagcaag ccatctgatt ttcatggtca gattgtaaag aagcaattac
     4141 tcttggacag tttaagcaga aaagccacca agtcaagtca ctggtcatat gatgaactcg
     4201 aagatctaat aaaagttaat ttaagcgttc attacaataa attcaagtga atctgtttta
     4261 agctagcctt taacgttttt aaattccttc aaacaaatta gttcctcttt gcagttcctc
     4321 atttctcggt gagctggtgc tttaatccct ttactttttc ttcctctagg gagtgatggt
     4381 tggcatggga cgaattaatt cctgcagccc gggggtaccg gtcgccacca tgaacacccc
     4441 gggaattaac ctgatcaagg aggacatgcg cgtgaaggtg cacatggagg gcaacgtgaa
     4501 cggccacgcc ttcgtgatcg agggcgaggg caagggcaag ccctacgagg gcacccagac
     4561 cgccaacctg accgtgaagg agggcgcccc cctgcccttc agctacgaca tcctgaccac
     4621 cgccgtgcac tacggcaacc gggtgttcac caagtacccc gaggacatcc ccgactactt
     4681 caagcagagc ttccccgagg gctacagctg ggagcgcacc atgaccttcg aggacaaggg
     4741 catctgcacc atccgcagcg acatcagcct ggagggcgac tgcttcttcc agaacgtgcg
     4801 cttcaagggc accaacttcc cccccaacgg ccccgtgatg cagaagaaaa ccctgaagtg
     4861 ggagcccagc accgagaagc tgcacgtgcg cgacggcctg ctggtgggca acatcaacat
     4921 ggccctgctg ctggagggcg gcggccacta cctgtgcgac ttcaagacca cctacaaggc
     4981 caagaaggtg gtgcagctgc ccgacgccca cttcgtggac caccgcatcg agatcctggg
     5041 caacgacagc gactacaaca aggtgaagct gtacgagcac gccgtggccc gctacagccc
     5101 cctgcccagc caggtgtggt aatctagatc ataatcagcc ataccacatt tgtagaggtt
     5161 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     5221 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     5281 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     5341 atcaatgtat cttaaggcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     5401 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcggat cctgcggccg
     5461 caagggaaaa tagaatgaag tgatctccaa aaaataagta ctttttgact gtaaataaaa
     5521 ttgtaaggag taaaaagtac ttttttttct aaaaaaatgt aattaagtaa aagtaaaagt
     5581 attgattttt aattgtactc aagtaaagta aaaatcccca aaaataatac ttaagtacag
     5641 taatcaagta aaattactca agtactttac acctctggtt cttgaccccc taccttcagc
     5701 aagcccagca gatccactag atcttttgtt ccctttagtg agggttaatt gcgcgcttgg
     5761 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     5821 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     5881 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     5941 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     6001 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     6061 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     6121 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     6181 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     6241 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     6301 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     6361 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     6421 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     6481 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     6541 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     6601 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     6661 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     6721 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     6781 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     6841 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     6901 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     6961 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     7021 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     7081 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     7141 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     7201 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     7261 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     7321 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     7381 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     7441 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     7501 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     7561 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     7621 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     7681 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     7741 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     7801 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     7861 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     7921 ctaaattgta agcgttaata t
//
