LOCUS       sequence-182554-        7597 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  empty vector for overexpression of microRNA and Dendra2 in 
            Zebrafish.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7597)
  AUTHORS   Hsu AY, Wang D, Gurol T, Zhou W, Zhu X, Lu HY, Deng Q
  TITLE     Overexpression of microRNA-722 fine-tunes neutrophilic inflammation 
            through inhibiting Rac2 in zebrafish.
  JOURNAL   Dis Model Mech. 2017 Sep 27. pii: dmm.030791. doi: 
            10.1242/dmm.030791.
  PUBMED    28954734
REFERENCE   2  (bases 1 to 7597)
  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: "Dis Model 
            Mech. 2017 Sep 27. pii: dmm.030791. doi: 10.1242/dmm.030791."
COMMENT     Sequence Label: Tol2-lyzC-Vector-Dendra2
FEATURES             Location/Qualifiers
     source          1..7597
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(159..180)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(576..587,588..725)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     regulatory      3153..3162
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4126..4818
                     /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    4929..5050
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4966..4985)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5020..5039
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    join(5174..5361,5362..5373)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5428..5448)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5428..5446)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5467..5483)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5467..5483)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5480..5502)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5491..5507
                     /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(5515..5521,5522..5539,5540..5545))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5560..5581
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5698..5715)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5869..6457)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5949..5968)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6628..7419,7420..7488))
                     /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     7251..7270
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7489..7593)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
       61 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
      121 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
      181 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
      241 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
      301 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gccacaatga
      361 cggctactac atggtgccat tccttcctct ttataggaat ggagactacc tcctgtccaa
      421 caatgctctt ggatacgagt acgcctacct gttggaccca ggtcattgca caacaccaga
      481 aatgccctct gatctgcaaa agacgtgaat atctgttcag acacccatat ccactctgtt
      541 ccacacaggt cagaggtttg tccaggagtt cttgacagag gtgtaaaaag tactcaaaaa
      601 ttttactcaa gtgaaagtac aagtacttag ggaaaatttt actcaattaa aagtaaaagt
      661 atctggctag aatcttactt gagtaaaagt aaaaaagtac tccattaaaa ttgtacttga
      721 gtattaagga agtaaaaggc tagcaagatc tgctcgagat acatttgatt agaagagttt
      781 gacttgggtt gattatagca aatagtggaa cacaaacaga ttttgaaata ttcttgcaaa
      841 tttatttatt aaatccattc aaatattgtc attttttgta ctgtgtaatt tatattgatt
      901 tgctcataca atatttacat ttcgcttaaa caatacacta ttatttatat ttaaaaatag
      961 ctatatataa atcaaatgta tagctctttt ttacacaatt aatcaaagct aaaatatttg
     1021 tcacacaaat acagtaagcc acttcacaat gatatcttca aaatctgtgc ttgactgaat
     1081 catttcattt attgttttgt aattcacatt ataccttacg ttctccaggc acagtttgaa
     1141 ttgttgataa atgaagataa tacatctgat ttaattagtc tcctttgttt ataataactt
     1201 taatctaatt tgatagtaat aattccagca atacatccat tacaataaaa atcgcttttt
     1261 tattttgcac attagtttgc cggtatttgt ttaaagtttc accatacttt taaatcatgt
     1321 gctatttgaa aagggctgtt atgactggaa ttaaatgttt ttttttatat cagtgtttta
     1381 gattgaagta caatatttat aggtaaaatt caccagcata tgcttaaaga tagccatgct
     1441 ccaggtcctt gtgtccttct aaggtctgtt tgtggtttaa gctgtttgtt gacacacgct
     1501 tgtcatctcc atttttctgt gaagttattg tcagtttatt gcacactggg ttgcattaag
     1561 ctgcagtttt ttcacaacag ttttccacat tgtaacagtc agtctccaaa ttattccatc
     1621 agtcctcaaa tttagtcatt tgaggccatg aaaacactta aaactacatg aaggtagttt
     1681 ggcattaggt tttagacttg ttcttgtttg gcatattgca acctgtgtaa aaatgatttg
     1741 atttttattt tttttttttt tttttcagaa agaaacggtc caaaaagagt tatgtgaaca
     1801 tttatccatg tgtttaatgt tttttaaatt aaaaaaaaac atttttatta aaccacttgt
     1861 gactaaacct tttttagtta ttctattttg tgacgagtct gaatggccaa agtactcttg
     1921 aatccaaaat ctttatagta aaatgtttaa tcttacttgc agtgttgggt ataatttgta
     1981 acaaagttga attccatatc tgctgagtga aaaagtaaag taagagattg ctgtaaatat
     2041 ttaggttatt tatttattaa tacaatatag aaagtaacaa aacacaatgt aaaaaatcaa
     2101 taaagaaagt aaatattaaa agtaatgata aactaatgat gtaaacaata ttagcaacaa
     2161 agtttgtaca attcagggag ggtaaagaat ttcacctaat agtcaaactc tgtattaaat
     2221 aatataaaat ctttgttctt agtctgaaga ggagactgtg agagaaagct tactggttgt
     2281 taaattagaa aatttgtaac ttgcagatta aaatgattta aataatttta taaaggccat
     2341 gtttgaaaca aatgtaaaaa cataataaat gttaagaatt tcagatattg agattttaca
     2401 atttaaaaaa agcaataaag taatagcaga gacctacact cttaaagtaa attacccaac
     2461 tctgcttact agtaactttt ggaatagtta ctagtatcca ttgaacaaat actagaatct
     2521 aattaaattg tgtgagtgtc caatgaaaag attcttgtat ttaaaacagt accagtatct
     2581 atatatgtac aaaatttagt gctagcaaaa tgcaaataaa tactactgca tataatggat
     2641 tgcattgtaa atcgactcgc cataatacat ttatttccta tttttacttt taacaaaaaa
     2701 taagttcact gagtatttca tgtcattaaa tcgcattaaa cctcttgcat gctttatgta
     2761 cactagtgca gttcgctaaa tgaccaccgg tgtcactaat aacacgatcg tttctgaatt
     2821 ctgcattaaa cgccatttat acgttgctct tattagtaaa caatttctcc atcatgagct
     2881 cgaccagtat tgcatatttc atcctcttaa atccagagtt gtttctttta atcactttct
     2941 cttcatgata aaagcagttt ttggttttca gaagtttaca acaaacccag tcagttattg
     3001 cctcagcaaa cactggagca actgcacact tgtgaaaagc ttcttgcgaa acacaaacct
     3061 ttctctggac ttataaataa aggcattcag cagctccacc atacacctcc actggtcttg
     3121 ataaagcaga tatcagcagt gatacaggta ccgccaccat ggatgaggaa atcgctgccc
     3181 tggtcgttga caacggctcc ggtatgtgca aagccggttt tgctggagat gatgcccctc
     3241 gtgctgtttt cccctccatt gttggacgac ccagacatca ggtacattta ataaaccatc
     3301 ctaataacat gttaatcata ttgaaccaca ttaaactctc atttgtgcat tttacatctt
     3361 ttaaaaccga tttatattcg tttttagata atgaagttta ttaaagtgtt aattcacagc
     3421 tttattgctt gttttgagaa tctgcttgca taagtgattt ttaaacttga tctgattttc
     3481 gtttgtttcg aagttcttca gctttgaaag tcagatcttc aattcacagc catgccatag
     3541 ttttaggata cagtataagt agacgaattg ggatgcatcg tagattttat gctgtgtttg
     3601 ttatggtttt tcagtggtag agattttcag agtattatta gggttatagt cagtgtcagg
     3661 tttatttatc gtcagcgtga tgggcaaaaa acagctggca gtacgggcta tgtcttcgtc
     3721 tagaagaaga caaagccaac tgctgttact ctcgcccaga aatctgcgcc aacatggaca
     3781 cacccagtgg agcaagccat ctgattttca tggtcagatt gtaaagaagc aattactctt
     3841 ggacagttta agcagaaaag ccaccaagtc aagtcactgg tcatatgatg aactcgaaga
     3901 tctaataaaa gttaatttaa gcgttcatta caataaattc aagtgaatct gttttaagct
     3961 agcctttaac gtttttaaat tccttcaaac aaattagttc ctctttgcag ttcctcattt
     4021 ctcggtgagc tggtgcttta atccctttac tttttcttcc tctagggagt gatggttggc
     4081 atgggacgaa ttaattcctg cagcccgggg gtaccggtcg ccaccatgaa caccccggga
     4141 attaacctga tcaaggagga catgcgcgtg aaggtgcaca tggagggcaa cgtgaacggc
     4201 cacgccttcg tgatcgaggg cgagggcaag ggcaagccct acgagggcac ccagaccgcc
     4261 aacctgaccg tgaaggaggg cgcccccctg cccttcagct acgacatcct gaccaccgcc
     4321 gtgcactacg gcaaccgggt gttcaccaag taccccgagg acatccccga ctacttcaag
     4381 cagagcttcc ccgagggcta cagctgggag cgcaccatga ccttcgagga caagggcatc
     4441 tgcaccatcc gcagcgacat cagcctggag ggcgactgct tcttccagaa cgtgcgcttc
     4501 aagggcacca acttcccccc caacggcccc gtgatgcaga agaaaaccct gaagtgggag
     4561 cccagcaccg agaagctgca cgtgcgcgac ggcctgctgg tgggcaacat caacatggcc
     4621 ctgctgctgg agggcggcgg ccactacctg tgcgacttca agaccaccta caaggccaag
     4681 aaggtggtgc agctgcccga cgcccacttc gtggaccacc gcatcgagat cctgggcaac
     4741 gacagcgact acaacaaggt gaagctgtac gagcacgccg tggcccgcta cagccccctg
     4801 cccagccagg tgtggtaatc tagatcataa tcagccatac cacatttgta gaggttttac
     4861 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     4921 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     4981 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     5041 atgtatctta aggcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     5101 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcggatcctg cggccgcaag
     5161 ggaaaataga atgaagtgat ctccaaaaaa taagtacttt ttgactgtaa ataaaattgt
     5221 aaggagtaaa aagtactttt ttttctaaaa aaatgtaatt aagtaaaagt aaaagtattg
     5281 atttttaatt gtactcaagt aaagtaaaaa tccccaaaaa taatacttaa gtacagtaat
     5341 caagtaaaat tactcaagta ctttacacct ctggttcttg accccctacc ttcagcaagc
     5401 ccagcagatc cactagatct tttgttccct ttagtgaggg ttaattgcgc gcttggcgta
     5461 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     5521 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     5581 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     5641 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     5701 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     5761 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5821 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5881 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5941 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     6001 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     6061 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     6121 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     6181 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     6241 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     6301 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     6361 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     6421 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     6481 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     6541 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     6601 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     6661 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     6721 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     6781 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     6841 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     6901 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     6961 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     7021 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     7081 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     7141 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     7201 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     7261 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     7321 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     7381 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     7441 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     7501 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     7561 tatttagaaa aataaacaaa taggggttcc gcgcaca
//
