LOCUS       sequence-182645-        7896 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 7896)
  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 7896)
  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 190a-Dendra2
FEATURES             Location/Qualifiers
     source          1..7896
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(111..132)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(528..539,540..677)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     regulatory      3105..3114
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4377..5069
                     /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    5180..5301
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5217..5236)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5271..5290
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    join(5425..5612,5613..5624)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5679..5699)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5679..5697)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5718..5734)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5718..5734)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5731..5753)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5742..5758
                     /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(5766..5772,5773..5790,5791..5796))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5811..5832
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5949..5966)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6120..6708)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6200..6219)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6879..7670,7671..7739))
                     /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     7502..7521
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7740..7844)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
       61 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
      121 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
      181 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
      241 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
      301 cggccacaat gacggctact acatggtgcc attccttcct ctttatagga atggagacta
      361 cctcctgtcc aacaatgctc ttggatacga gtacgcctac ctgttggacc caggtcattg
      421 cacaacacca gaaatgccct ctgatctgca aaagacgtga atatctgttc agacacccat
      481 atccactctg ttccacacag gtcagaggtt tgtccaggag ttcttgacag aggtgtaaaa
      541 agtactcaaa aattttactc aagtgaaagt acaagtactt agggaaaatt ttactcaatt
      601 aaaagtaaaa gtatctggct agaatcttac ttgagtaaaa gtaaaaaagt actccattaa
      661 aattgtactt gagtattaag gaagtaaaag gctagcaaga tctgctcgag atacatttga
      721 ttagaagagt ttgacttggg ttgattatag caaatagtgg aacacaaaca gattttgaaa
      781 tattcttgca aatttattta ttaaatccat tcaaatattg tcattttttg tactgtgtaa
      841 tttatattga tttgctcata caatatttac atttcgctta aacaatacac tattatttat
      901 atttaaaaat agctatatat aaatcaaatg tatagctctt ttttacacaa ttaatcaaag
      961 ctaaaatatt tgtcacacaa atacagtaag ccacttcaca atgatatctt caaaatctgt
     1021 gcttgactga atcatttcat ttattgtttt gtaattcaca ttatacctta cgttctccag
     1081 gcacagtttg aattgttgat aaatgaagat aatacatctg atttaattag tctcctttgt
     1141 ttataataac tttaatctaa tttgatagta ataattccag caatacatcc attacaataa
     1201 aaatcgcttt tttattttgc acattagttt gccggtattt gtttaaagtt tcaccatact
     1261 tttaaatcat gtgctatttg aaaagggctg ttatgactgg aattaaatgt ttttttttat
     1321 atcagtgttt tagattgaag tacaatattt ataggtaaaa ttcaccagca tatgcttaaa
     1381 gatagccatg ctccaggtcc ttgtgtcctt ctaaggtctg tttgtggttt aagctgtttg
     1441 ttgacacacg cttgtcatct ccatttttct gtgaagttat tgtcagttta ttgcacactg
     1501 ggttgcatta agctgcagtt ttttcacaac agttttccac attgtaacag tcagtctcca
     1561 aattattcca tcagtcctca aatttagtca tttgaggcca tgaaaacact taaaactaca
     1621 tgaaggtagt ttggcattag gttttagact tgttcttgtt tggcatattg caacctgtgt
     1681 aaaaatgatt tgatttttat tttttttttt tttttttcag aaagaaacgg tccaaaaaga
     1741 gttatgtgaa catttatcca tgtgtttaat gttttttaaa ttaaaaaaaa acatttttat
     1801 taaaccactt gtgactaaac cttttttagt tattctattt tgtgacgagt ctgaatggcc
     1861 aaagtactct tgaatccaaa atctttatag taaaatgttt aatcttactt gcagtgttgg
     1921 gtataatttg taacaaagtt gaattccata tctgctgagt gaaaaagtaa agtaagagat
     1981 tgctgtaaat atttaggtta tttatttatt aatacaatat agaaagtaac aaaacacaat
     2041 gtaaaaaatc aataaagaaa gtaaatatta aaagtaatga taaactaatg atgtaaacaa
     2101 tattagcaac aaagtttgta caattcaggg agggtaaaga atttcaccta atagtcaaac
     2161 tctgtattaa ataatataaa atctttgttc ttagtctgaa gaggagactg tgagagaaag
     2221 cttactggtt gttaaattag aaaatttgta acttgcagat taaaatgatt taaataattt
     2281 tataaaggcc atgtttgaaa caaatgtaaa aacataataa atgttaagaa tttcagatat
     2341 tgagatttta caatttaaaa aaagcaataa agtaatagca gagacctaca ctcttaaagt
     2401 aaattaccca actctgctta ctagtaactt ttggaatagt tactagtatc cattgaacaa
     2461 atactagaat ctaattaaat tgtgtgagtg tccaatgaaa agattcttgt atttaaaaca
     2521 gtaccagtat ctatatatgt acaaaattta gtgctagcaa aatgcaaata aatactactg
     2581 catataatgg attgcattgt aaatcgactc gccataatac atttatttcc tatttttact
     2641 tttaacaaaa aataagttca ctgagtattt catgtcatta aatcgcatta aacctcttgc
     2701 atgctttatg tacactagtg cagttcgcta aatgaccacc ggtgtcacta ataacacgat
     2761 cgtttctgaa ttctgcatta aacgccattt atacgttgct cttattagta aacaatttct
     2821 ccatcatgag ctcgaccagt attgcatatt tcatcctctt aaatccagag ttgtttcttt
     2881 taatcacttt ctcttcatga taaaagcagt ttttggtttt cagaagttta caacaaaccc
     2941 agtcagttat tgcctcagca aacactggag caactgcaca cttgtgaaaa gcttcttgcg
     3001 aaacacaaac ctttctctgg acttataaat aaaggcattc agcagctcca ccatacacct
     3061 ccactggtct tgataaagca gatatcagca gtgatacagg taccgccacc atggatgagg
     3121 aaatcgctgc cctggtcgtt gacaacggct ccggtatgtg caaagccggt tttgctggag
     3181 atgatgcccc tcgtgctgtt ttcccctcca ttgttggacg acccagacat caggtacatt
     3241 taataaacca tcctaataac atgttaatca tattgaacca cattaaactc tcatttgtgc
     3301 attttacatc ttttaaaacc gatttatatt cgtttttaga taatgaagtt tattaaagtg
     3361 ttaattcaca gctttattgc ttgttttgag aatctgcttg cataagtgat ttttaaactt
     3421 gatctgattt tcgtttgttt cgaagttctt cagctttgaa agtcagatct tcaattcaca
     3481 gccatgccat agttttagga tacagtataa gtagacgaat tgggatgcat cgtagatttt
     3541 atgctgtgtt tgttatggtt tttcagtggt agagattttc agagtattat tagggttata
     3601 gtcagtgtca ggtttattta tcgtcagcgt gatgggcaaa aaacagctgg cagtacgggc
     3661 tgcctctcct cctctaccca tcatgttttt tccttcgtta tcccagcatg ccctgctctc
     3721 ctctcccctc ccccgtgttc tggaggtgag gtagacctgg aagcctttct gcaggcctct
     3781 gtttgatatg tttgatatat taggttgtta ttctgtccaa ctatatatca aacatattcc
     3841 tacagtgtcc tgctctgtct ccagactcgt gtgtgttttg ttcatgaacg tgtgtgtgtg
     3901 tgtgtgtgtg tgtgtgtgtg gcctcagaca gatcagtttt gtgaaggtgg cgtgcaaggc
     3961 tttgtcttag gcccactgtg cttgagccaa ctgctgttac tctcgcccag aaatctgcgc
     4021 caacatggac acacccagtg gagcaagcca tctgattttc atggtcagat tgtaaagaag
     4081 caattactct tggacagttt aagcagaaaa gccaccaagt caagtcactg gtcatatgat
     4141 gaactcgaag atctaataaa agttaattta agcgttcatt acaataaatt caagtgaatc
     4201 tgttttaagc tagcctttaa cgtttttaaa ttccttcaaa caaattagtt cctctttgca
     4261 gttcctcatt tctcggtgag ctggtgcttt aatcccttta ctttttcttc ctctagggag
     4321 tgatggttgg catgggacga attaattcct gcagcccggg ggtaccggtc gccaccatga
     4381 acaccccggg aattaacctg atcaaggagg acatgcgcgt gaaggtgcac atggagggca
     4441 acgtgaacgg ccacgccttc gtgatcgagg gcgagggcaa gggcaagccc tacgagggca
     4501 cccagaccgc caacctgacc gtgaaggagg gcgcccccct gcccttcagc tacgacatcc
     4561 tgaccaccgc cgtgcactac ggcaaccggg tgttcaccaa gtaccccgag gacatccccg
     4621 actacttcaa gcagagcttc cccgagggct acagctggga gcgcaccatg accttcgagg
     4681 acaagggcat ctgcaccatc cgcagcgaca tcagcctgga gggcgactgc ttcttccaga
     4741 acgtgcgctt caagggcacc aacttccccc ccaacggccc cgtgatgcag aagaaaaccc
     4801 tgaagtggga gcccagcacc gagaagctgc acgtgcgcga cggcctgctg gtgggcaaca
     4861 tcaacatggc cctgctgctg gagggcggcg gccactacct gtgcgacttc aagaccacct
     4921 acaaggccaa gaaggtggtg cagctgcccg acgcccactt cgtggaccac cgcatcgaga
     4981 tcctgggcaa cgacagcgac tacaacaagg tgaagctgta cgagcacgcc gtggcccgct
     5041 acagccccct gcccagccag gtgtggtaat ctagatcata atcagccata ccacatttgt
     5101 agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat
     5161 gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa
     5221 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5281 caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg ttaaaattcg
     5341 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcggatcct
     5401 gcggccgcaa gggaaaatag aatgaagtga tctccaaaaa ataagtactt tttgactgta
     5461 aataaaattg taaggagtaa aaagtacttt tttttctaaa aaaatgtaat taagtaaaag
     5521 taaaagtatt gatttttaat tgtactcaag taaagtaaaa atccccaaaa ataatactta
     5581 agtacagtaa tcaagtaaaa ttactcaagt actttacacc tctggttctt gaccccctac
     5641 cttcagcaag cccagcagat ccactagatc ttttgttccc tttagtgagg gttaattgcg
     5701 cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     5761 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5821 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     5881 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     5941 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     6001 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     6061 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     6121 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     6181 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     6241 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     6301 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     6361 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     6421 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     6481 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     6541 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     6601 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     6661 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     6721 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     6781 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     6841 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     6901 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     6961 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     7021 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     7081 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     7141 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     7201 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     7261 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     7321 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     7381 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     7441 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     7501 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     7561 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     7621 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     7681 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     7741 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     7801 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     7861 gtgccaccta aattgtaagc gttaatattt tgttaa
//
