LOCUS       sequence-182923-        6654 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For dual luciferase report assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6654)
  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 6654)
  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: pSi-check2-zRac2 3'UTR
FEATURES             Location/Qualifiers
     source          1..6654
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      9..597
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     498..517
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        726..1083
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        726..917
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(749..769)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      934..1069
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          1153..1285
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1268..1287
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1330..1348
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1352..1361
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1358..2293
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    2733..2781
                     /note="synthetic polyadenylation signal"
     promoter        2790..3541
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      3571..3580
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3577..5229
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /db_xref="GI:42718124"
                     /protein_id="AAS38485.1"
                     /translation="MADAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     RBS             4536..4544
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    5273..5394
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5284..5303)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5338..5357
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5527..5631
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5632..5700,5701..6492)
                     /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(5850..5869)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
       61 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
      121 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
      181 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
      241 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
      301 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
      361 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
      421 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
      481 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
      541 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
      601 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatggct
      661 cgacagatct gcgcagcacc atggcctgaa ataacctctg aaagaggaac ttggttaggt
      721 accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
      781 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
      841 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
      901 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
      961 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     1021 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     1081 aaaaagcttg attcttctga cacaacagtc tcgaacttaa gctgcagaag ttggtcgtga
     1141 ggcactgggc aggtaagtat caaggttaca agacaggttt aaggagacca atagaaactg
     1201 ggcttgtcga gacagagaag actcttgcgt ttctgatagg cacctattgg tcttactgac
     1261 atccactttg cctttctctc cacaggtgtc cactcccagt tcaattacag ctcttaaggc
     1321 tagagtactt aatacgactc actataggct agccaccatg gcttccaagg tgtacgaccc
     1381 cgagcaacgc aaacgcatga tcactgggcc tcagtggtgg gctcgctgca agcaaatgaa
     1441 cgtgctggac tccttcatca actactatga ttccgagaag cacgccgaga acgccgtgat
     1501 ttttctgcat ggtaacgctg cctccagcta cctgtggagg cacgtcgtgc ctcacatcga
     1561 gcccgtggct agatgcatca tccctgatct gatcggaatg ggtaagtccg gcaagagcgg
     1621 gaatggctca tatcgcctcc tggatcacta caagtacctc accgcttggt tcgagctgct
     1681 gaaccttcca aagaaaatca tctttgtggg ccacgactgg ggggcttgtc tggcctttca
     1741 ctactcctac gagcaccaag acaagatcaa ggccatcgtc catgctgaga gtgtcgtgga
     1801 cgtgatcgag tcctgggacg agtggcctga catcgaggag gatatcgccc tgatcaagag
     1861 cgaagagggc gagaaaatgg tgcttgagaa taacttcttc gtcgagacca tgctcccaag
     1921 caagatcatg cggaaactgg agcctgagga gttcgctgcc tacctggagc cattcaagga
     1981 gaagggcgag gttagacggc ctaccctctc ctggcctcgc gagatccctc tcgttaaggg
     2041 aggcaagccc gacgtcgtcc agattgtccg caactacaac gcctaccttc gggccagcga
     2101 cgatctgcct aagatgttca tcgagtccga ccctgggttc ttttccaacg ctattgtcga
     2161 gggagctaag aagttcccta acaccgagtt cgtgaaggtg aagggcctcc acttcagcca
     2221 ggaggacgct ccagatgaaa tgggtaagta catcaagagc ttcgtggagc gcgtgctgaa
     2281 gaacgagcag taattctagg cgatcgctcg agagatacac gattcgtcac tgtaaaaaag
     2341 aaaagggtac cagaggaatc aactggattt ctcctataaa ccaacacaac atgcaattgc
     2401 cttacgaaag cttttcctca atttttagct gaatatatct tttgaacatg ccacattgat
     2461 gccgatacag tgataatttt agtaacagaa tatttaatca tgttttaact gatgtgttat
     2521 gttattttga gactgtagtg tgtttgtcag gtacgcagtt gcatattgtt ttctagcgac
     2581 aacataaaca gctgaatttt ttattttttt gttacacata attgaaccca caggaaggta
     2641 ttagacatga atcatattca tgttatacgt atggaattat ttttggcaga aaatggcaaa
     2701 aataaactgt acaactggcg gccgctggcc gcaataaaat atctttattt tcattacatc
     2761 tgtgtgttgg ttttttgtgt gaggatctaa atgagtcttc ggacctcgcg ggggccgctt
     2821 aagcggtggt tagggtttgt ctgacgcggg gggaggggga aggaacgaaa cactctcatt
     2881 cggaggcggc tcggggtttg gtcttggtgg ccacgggcac gcagaagagc gccgcgatcc
     2941 tcttaagcac ccccccgccc tccgtggagg cgggggtttg gtcggcgggt ggtaactggc
     3001 gggccgctga ctcgggcggg tcgcgcgccc cagagtgtga ccttttcggt ctgctcgcag
     3061 acccccgggc ggcgccgccg cggcggcgac gggctcgctg ggtcctaggc tccatgggga
     3121 ccgtatacgt ggacaggctc tggagcatcc gcacgactgc ggtgatatta ccggagacct
     3181 tctgcgggac gagccgggtc acgcggctga cgcggagcgt ccgttgggcg acaaacacca
     3241 ggacggggca caggtacact atcttgtcac ccggaggcgc gagggactgc aggagcttca
     3301 gggagtggcg cagctgcttc atccccgtgg cccgttgctc gcgtttgctg gcggtgtccc
     3361 cggaagaaat atatttgcat gtctttagtt ctatgatgac acaaaccccg cccagcgtct
     3421 tgtcattggc gaattcgaac acgcagatgc agtcggggcg gcgcggtccc aggtccactt
     3481 cgcatattaa ggtgacgcgt gtggcctcga acaccgagcg accctgcagc gacccgctta
     3541 aaagcttggc attccggtac tgttggtaaa gccaccatgg ccgatgctaa gaacattaag
     3601 aagggccctg ctcccttcta ccctctggag gatggcaccg ctggcgagca gctgcacaag
     3661 gccatgaaga ggtatgccct ggtgcctggc accattgcct tcaccgatgc ccacattgag
     3721 gtggacatca cctatgccga gtacttcgag atgtctgtgc gcctggccga ggccatgaag
     3781 aggtacggcc tgaacaccaa ccaccgcatc gtggtgtgct ctgagaactc tctgcagttc
     3841 ttcatgccag tgctgggcgc cctgttcatc ggagtggccg tggcccctgc taacgacatt
     3901 tacaacgagc gcgagctgct gaacagcatg ggcatttctc agcctaccgt ggtgttcgtg
     3961 tctaagaagg gcctgcagaa gatcctgaac gtgcagaaga agctgcctat catccagaag
     4021 atcatcatca tggactctaa gaccgactac cagggcttcc agagcatgta cacattcgtg
     4081 acatctcatc tgcctcctgg cttcaacgag tacgacttcg tgccagagtc tttcgacagg
     4141 gacaaaacca ttgccctgat catgaacagc tctgggtcta ccggcctgcc taagggcgtg
     4201 gccctgcctc atcgcaccgc ctgtgtgcgc ttctctcacg cccgcgaccc tattttcggc
     4261 aaccagatca tccccgacac cgctattctg agcgtggtgc cattccacca cggcttcggc
     4321 atgttcacca ccctgggcta cctgatttgc ggctttcggg tggtgctgat gtaccgcttc
     4381 gaggaggagc tgttcctgcg cagcctgcaa gactacaaaa ttcagtctgc cctgctggtg
     4441 ccaaccctgt tcagcttctt cgctaagagc accctgatcg acaagtacga cctgtctaac
     4501 ctgcacgaga ttgcctctgg cggcgcccca ctgtctaagg aggtgggcga agccgtggcc
     4561 aagcgctttc atctgccagg catccgccag ggctacggcc tgaccgagac aaccagcgcc
     4621 attctgatta ccccagaggg cgacgacaag cctggcgccg tgggcaaggt ggtgccattc
     4681 ttcgaggcca aggtggtgga cctggacacc ggcaagaccc tgggagtgaa ccagcgcggc
     4741 gagctgtgtg tgcgcggccc tatgattatg tccggctacg tgaataaccc tgaggccaca
     4801 aacgccctga tcgacaagga cggctggctg cactctggcg acattgccta ctgggacgag
     4861 gacgagcact tcttcatcgt ggaccgcctg aagtctctga tcaagtacaa gggctaccag
     4921 gtggccccag ccgagctgga gtctatcctg ctgcagcacc ctaacatttt cgacgccgga
     4981 gtggccggcc tgcccgacga cgatgccggc gagctgcctg ccgccgtcgt cgtgctggaa
     5041 cacggcaaga ccatgaccga gaaggagatc gtggactatg tggccagcca ggtgacaacc
     5101 gccaagaagc tgcgcggcgg agtggtgttc gtggacgagg tgcccaaggg cctgaccggc
     5161 aagctggacg cccgcaagat ccgcgagatc ctgatcaagg ctaagaaagg cggcaagatc
     5221 gccgtgtaat aattctagag tcggggcggc cggccgcttc gagcagacat gataagatac
     5281 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     5341 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     5401 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc
     5461 aagtaaaacc tctacaaatg tggtaaaatc gataaggatc caggtggcac ttttcgggga
     5521 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     5581 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     5641 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     5701 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     5761 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     5821 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     5881 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     5941 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     6001 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     6061 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     6121 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     6181 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     6241 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     6301 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     6361 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     6421 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     6481 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     6541 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     6601 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaa
//
