LOCUS       sequence-204470-        7859 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 7859)
  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 7859)
  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-miR-23a-2-dendra2
FEATURES             Location/Qualifiers
     source          1..7859
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(144..165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(561..572,573..710)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     regulatory      3138..3147
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4373..5065
                     /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    5176..5297
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5213..5232)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5267..5286
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    join(5421..5608,5609..5620)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5675..5695)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5675..5693)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5714..5730)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5714..5730)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5727..5749)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5738..5754
                     /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(5762..5768,5769..5786,5787..5792))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5807..5828
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5945..5962)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6116..6704)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6196..6215)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6875..7666,7667..7735))
                     /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     7498..7517
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7736..7840)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
       61 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
      121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
      181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
      241 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
      301 ggagcccccg atttagagct tgacggggaa agccggccac aatgacggct actacatggt
      361 gccattcctt cctctttata ggaatggaga ctacctcctg tccaacaatg ctcttggata
      421 cgagtacgcc tacctgttgg acccaggtca ttgcacaaca ccagaaatgc cctctgatct
      481 gcaaaagacg tgaatatctg ttcagacacc catatccact ctgttccaca caggtcagag
      541 gtttgtccag gagttcttga cagaggtgta aaaagtactc aaaaatttta ctcaagtgaa
      601 agtacaagta cttagggaaa attttactca attaaaagta aaagtatctg gctagaatct
      661 tacttgagta aaagtaaaaa agtactccat taaaattgta cttgagtatt aaggaagtaa
      721 aaggctagca agatctgctc gagatacatt tgattagaag agtttgactt gggttgatta
      781 tagcaaatag tggaacacaa acagattttg aaatattctt gcaaatttat ttattaaatc
      841 cattcaaata ttgtcatttt ttgtactgtg taatttatat tgatttgctc atacaatatt
      901 tacatttcgc ttaaacaata cactattatt tatatttaaa aatagctata tataaatcaa
      961 atgtatagct cttttttaca caattaatca aagctaaaat atttgtcaca caaatacagt
     1021 aagccacttc acaatgatat cttcaaaatc tgtgcttgac tgaatcattt catttattgt
     1081 tttgtaattc acattatacc ttacgttctc caggcacagt ttgaattgtt gataaatgaa
     1141 gataatacat ctgatttaat tagtctcctt tgtttataat aactttaatc taatttgata
     1201 gtaataattc cagcaataca tccattacaa taaaaatcgc ttttttattt tgcacattag
     1261 tttgccggta tttgtttaaa gtttcaccat acttttaaat catgtgctat ttgaaaaggg
     1321 ctgttatgac tggaattaaa tgtttttttt tatatcagtg ttttagattg aagtacaata
     1381 tttataggta aaattcacca gcatatgctt aaagatagcc atgctccagg tccttgtgtc
     1441 cttctaaggt ctgtttgtgg tttaagctgt ttgttgacac acgcttgtca tctccatttt
     1501 tctgtgaagt tattgtcagt ttattgcaca ctgggttgca ttaagctgca gttttttcac
     1561 aacagttttc cacattgtaa cagtcagtct ccaaattatt ccatcagtcc tcaaatttag
     1621 tcatttgagg ccatgaaaac acttaaaact acatgaaggt agtttggcat taggttttag
     1681 acttgttctt gtttggcata ttgcaacctg tgtaaaaatg atttgatttt tatttttttt
     1741 tttttttttt cagaaagaaa cggtccaaaa agagttatgt gaacatttat ccatgtgttt
     1801 aatgtttttt aaattaaaaa aaaacatttt tattaaacca cttgtgacta aacctttttt
     1861 agttattcta ttttgtgacg agtctgaatg gccaaagtac tcttgaatcc aaaatcttta
     1921 tagtaaaatg tttaatctta cttgcagtgt tgggtataat ttgtaacaaa gttgaattcc
     1981 atatctgctg agtgaaaaag taaagtaaga gattgctgta aatatttagg ttatttattt
     2041 attaatacaa tatagaaagt aacaaaacac aatgtaaaaa atcaataaag aaagtaaata
     2101 ttaaaagtaa tgataaacta atgatgtaaa caatattagc aacaaagttt gtacaattca
     2161 gggagggtaa agaatttcac ctaatagtca aactctgtat taaataatat aaaatctttg
     2221 ttcttagtct gaagaggaga ctgtgagaga aagcttactg gttgttaaat tagaaaattt
     2281 gtaacttgca gattaaaatg atttaaataa ttttataaag gccatgtttg aaacaaatgt
     2341 aaaaacataa taaatgttaa gaatttcaga tattgagatt ttacaattta aaaaaagcaa
     2401 taaagtaata gcagagacct acactcttaa agtaaattac ccaactctgc ttactagtaa
     2461 cttttggaat agttactagt atccattgaa caaatactag aatctaatta aattgtgtga
     2521 gtgtccaatg aaaagattct tgtatttaaa acagtaccag tatctatata tgtacaaaat
     2581 ttagtgctag caaaatgcaa ataaatacta ctgcatataa tggattgcat tgtaaatcga
     2641 ctcgccataa tacatttatt tcctattttt acttttaaca aaaaataagt tcactgagta
     2701 tttcatgtca ttaaatcgca ttaaacctct tgcatgcttt atgtacacta gtgcagttcg
     2761 ctaaatgacc accggtgtca ctaataacac gatcgtttct gaattctgca ttaaacgcca
     2821 tttatacgtt gctcttatta gtaaacaatt tctccatcat gagctcgacc agtattgcat
     2881 atttcatcct cttaaatcca gagttgtttc ttttaatcac tttctcttca tgataaaagc
     2941 agtttttggt tttcagaagt ttacaacaaa cccagtcagt tattgcctca gcaaacactg
     3001 gagcaactgc acacttgtga aaagcttctt gcgaaacaca aacctttctc tggacttata
     3061 aataaaggca ttcagcagct ccaccataca cctccactgg tcttgataaa gcagatatca
     3121 gcagtgatac aggtaccgcc accatggatg aggaaatcgc tgccctggtc gttgacaacg
     3181 gctccggtat gtgcaaagcc ggttttgctg gagatgatgc ccctcgtgct gttttcccct
     3241 ccattgttgg acgacccaga catcaggtac atttaataaa ccatcctaat aacatgttaa
     3301 tcatattgaa ccacattaaa ctctcatttg tgcattttac atcttttaaa accgatttat
     3361 attcgttttt agataatgaa gtttattaaa gtgttaattc acagctttat tgcttgtttt
     3421 gagaatctgc ttgcataagt gatttttaaa cttgatctga ttttcgtttg tttcgaagtt
     3481 cttcagcttt gaaagtcaga tcttcaattc acagccatgc catagtttta ggatacagta
     3541 taagtagacg aattgggatg catcgtagat tttatgctgt gtttgttatg gtttttcagt
     3601 ggtagagatt ttcagagtat tattagggtt atagtcagtg tcaggtttat ttatcgtcag
     3661 cgtgatgggc aaaaaacagc tggcagtacg ggctatatag ccgtgtcggt ccctcagaca
     3721 cagcgactta aatccagacc ctcctcttct gccggccagg ggaattcctg gcagagtgat
     3781 ttttaaacct aatgactgaa tcacattgcc agggatttcc aatggctcgt gtgactcaga
     3841 gacacgccag atataaatac tcggcaaaca cacacacacc tgaacacatg agaagctaat
     3901 cctaaaggaa tgccgactgt ctcacacact agttagcggt gtcaggaatt catgtgcact
     3961 tctatgcggg ttgtgtggtg agccaactgc tgttactctc gcccagaaat ctgcgccaac
     4021 atggacacac ccagtggagc aagccatctg attttcatgg tcagattgta aagaagcaat
     4081 tactcttgga cagtttaagc agaaaagcca ccaagtcaag tcactggtca tatgatgaac
     4141 tcgaagatct aataaaagtt aatttaagcg ttcattacaa taaattcaag tgaatctgtt
     4201 ttaagctagc ctttaacgtt tttaaattcc ttcaaacaaa ttagttcctc tttgcagttc
     4261 ctcatttctc ggtgagctgg tgctttaatc cctttacttt ttcttcctct agggagtgat
     4321 ggttggcatg ggacgaatta attcctgcag cccgggggta ccggtcgcca ccatgaacac
     4381 cccgggaatt aacctgatca aggaggacat gcgcgtgaag gtgcacatgg agggcaacgt
     4441 gaacggccac gccttcgtga tcgagggcga gggcaagggc aagccctacg agggcaccca
     4501 gaccgccaac ctgaccgtga aggagggcgc ccccctgccc ttcagctacg acatcctgac
     4561 caccgccgtg cactacggca accgggtgtt caccaagtac cccgaggaca tccccgacta
     4621 cttcaagcag agcttccccg agggctacag ctgggagcgc accatgacct tcgaggacaa
     4681 gggcatctgc accatccgca gcgacatcag cctggagggc gactgcttct tccagaacgt
     4741 gcgcttcaag ggcaccaact tcccccccaa cggccccgtg atgcagaaga aaaccctgaa
     4801 gtgggagccc agcaccgaga agctgcacgt gcgcgacggc ctgctggtgg gcaacatcaa
     4861 catggccctg ctgctggagg gcggcggcca ctacctgtgc gacttcaaga ccacctacaa
     4921 ggccaagaag gtggtgcagc tgcccgacgc ccacttcgtg gaccaccgca tcgagatcct
     4981 gggcaacgac agcgactaca acaaggtgaa gctgtacgag cacgccgtgg cccgctacag
     5041 ccccctgccc agccaggtgt ggtaatctag atcataatca gccataccac atttgtagag
     5101 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     5161 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     5221 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     5281 ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt
     5341 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggcg gatcctgcgg
     5401 ccgcaaggga aaatagaatg aagtgatctc caaaaaataa gtactttttg actgtaaata
     5461 aaattgtaag gagtaaaaag tacttttttt tctaaaaaaa tgtaattaag taaaagtaaa
     5521 agtattgatt tttaattgta ctcaagtaaa gtaaaaatcc ccaaaaataa tacttaagta
     5581 cagtaatcaa gtaaaattac tcaagtactt tacacctctg gttcttgacc ccctaccttc
     5641 agcaagccca gcagatccac tagatctttt gttcccttta gtgagggtta attgcgcgct
     5701 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     5761 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     5821 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     5881 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     5941 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     6001 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     6061 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     6121 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     6181 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     6241 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     6301 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     6361 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     6421 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     6481 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     6541 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     6601 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     6661 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     6721 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     6781 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6841 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6901 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6961 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     7021 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     7081 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     7141 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     7201 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     7261 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     7321 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     7381 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     7441 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     7501 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     7561 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     7621 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     7681 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     7741 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     7801 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtg
//
