LOCUS       sequence-230279-        6792 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  luciferase assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6792)
  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 6792)
  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: dre-cdk2 3'UTR sicheck2 199 mut
FEATURES             Location/Qualifiers
     source          1..6792
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        62..419
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        62..253
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(85..105)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      270..405
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          489..621
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     604..623
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        666..684
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      688..697
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             694..1629
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    2207..2255
                     /note="synthetic polyadenylation signal"
     promoter        2264..3015
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      3045..3054
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3051..4703
                     /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             4010..4018
                     /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    4747..4868
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4758..4777)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4812..4831
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5001..5105
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5106..5174,5175..5966)
                     /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(5324..5343)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6137..6725
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6626..6645
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agatctgcgc agcaccatgg cctgaaataa cctctgaaag aggaacttgg ttaggtacct
       61 tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
      121 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
      181 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
      241 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
      301 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
      361 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
      421 agcttgattc ttctgacaca acagtctcga acttaagctg cagaagttgg tcgtgaggca
      481 ctgggcaggt aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct
      541 tgtcgagaca gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc
      601 actttgcctt tctctccaca ggtgtccact cccagttcaa ttacagctct taaggctaga
      661 gtacttaata cgactcacta taggctagcc accatggctt ccaaggtgta cgaccccgag
      721 caacgcaaac gcatgatcac tgggcctcag tggtgggctc gctgcaagca aatgaacgtg
      781 ctggactcct tcatcaacta ctatgattcc gagaagcacg ccgagaacgc cgtgattttt
      841 ctgcatggta acgctgcctc cagctacctg tggaggcacg tcgtgcctca catcgagccc
      901 gtggctagat gcatcatccc tgatctgatc ggaatgggta agtccggcaa gagcgggaat
      961 ggctcatatc gcctcctgga tcactacaag tacctcaccg cttggttcga gctgctgaac
     1021 cttccaaaga aaatcatctt tgtgggccac gactgggggg cttgtctggc ctttcactac
     1081 tcctacgagc accaagacaa gatcaaggcc atcgtccatg ctgagagtgt cgtggacgtg
     1141 atcgagtcct gggacgagtg gcctgacatc gaggaggata tcgccctgat caagagcgaa
     1201 gagggcgaga aaatggtgct tgagaataac ttcttcgtcg agaccatgct cccaagcaag
     1261 atcatgcgga aactggagcc tgaggagttc gctgcctacc tggagccatt caaggagaag
     1321 ggcgaggtta gacggcctac cctctcctgg cctcgcgaga tccctctcgt taagggaggc
     1381 aagcccgacg tcgtccagat tgtccgcaac tacaacgcct accttcgggc cagcgacgat
     1441 ctgcctaaga tgttcatcga gtccgaccct gggttctttt ccaacgctat tgtcgaggga
     1501 gctaagaagt tccctaacac cgagttcgtg aaggtgaagg gcctccactt cagccaggag
     1561 gacgctccag atgaaatggg taagtacatc aagagcttcg tggagcgcgt gctgaagaac
     1621 gagcagtaat tctaggcgat cgctcgagaa cgagatcaac tttggcaaga cgggactaaa
     1681 agtttcacac aaaacttttc tgaaaatgat ttcccagttt ggaaatcgtt cggttttata
     1741 atcgatttgt acgctcgttt tcctaaagtc actctgggtt tgttttaatc tgaagattat
     1801 ccattgcaac ctgtgtgacc caaaaaaaca gctgtgctaa tagtatgcgt gtgattaaaa
     1861 tgactcggag gatgtaaatt tgcataaagc agacgtgatg ttactaacat aactttagtt
     1921 cggagatgac attccatggt tatattgctg atataacttg tgacgttgta ttgcttatat
     1981 cacacaagta catatcgcat tgagcgcttc gtctagcctt agactgaatg aaactgttta
     2041 gactccctgt atctttttgc tgctttattc caagattata ttattataat taaagatttt
     2101 aaacctaatt tgcttttaaa gtttcctaaa aatagttctc tctgcatctt catccattaa
     2161 actcgtataa catttggttt atgttgttac agcggccgct ggccgcaata aaatatcttt
     2221 attttcatta catctgtgtg ttggtttttt gtgtgaggat ctaaatgagt cttcggacct
     2281 cgcgggggcc gcttaagcgg tggttagggt ttgtctgacg cggggggagg gggaaggaac
     2341 gaaacactct cattcggagg cggctcgggg tttggtcttg gtggccacgg gcacgcagaa
     2401 gagcgccgcg atcctcttaa gcaccccccc gccctccgtg gaggcggggg tttggtcggc
     2461 gggtggtaac tggcgggccg ctgactcggg cgggtcgcgc gccccagagt gtgacctttt
     2521 cggtctgctc gcagaccccc gggcggcgcc gccgcggcgg cgacgggctc gctgggtcct
     2581 aggctccatg gggaccgtat acgtggacag gctctggagc atccgcacga ctgcggtgat
     2641 attaccggag accttctgcg ggacgagccg ggtcacgcgg ctgacgcgga gcgtccgttg
     2701 ggcgacaaac accaggacgg ggcacaggta cactatcttg tcacccggag gcgcgaggga
     2761 ctgcaggagc ttcagggagt ggcgcagctg cttcatcccc gtggcccgtt gctcgcgttt
     2821 gctggcggtg tccccggaag aaatatattt gcatgtcttt agttctatga tgacacaaac
     2881 cccgcccagc gtcttgtcat tggcgaattc gaacacgcag atgcagtcgg ggcggcgcgg
     2941 tcccaggtcc acttcgcata ttaaggtgac gcgtgtggcc tcgaacaccg agcgaccctg
     3001 cagcgacccg cttaaaagct tggcattccg gtactgttgg taaagccacc atggccgatg
     3061 ctaagaacat taagaagggc cctgctccct tctaccctct ggaggatggc accgctggcg
     3121 agcagctgca caaggccatg aagaggtatg ccctggtgcc tggcaccatt gccttcaccg
     3181 atgcccacat tgaggtggac atcacctatg ccgagtactt cgagatgtct gtgcgcctgg
     3241 ccgaggccat gaagaggtac ggcctgaaca ccaaccaccg catcgtggtg tgctctgaga
     3301 actctctgca gttcttcatg ccagtgctgg gcgccctgtt catcggagtg gccgtggccc
     3361 ctgctaacga catttacaac gagcgcgagc tgctgaacag catgggcatt tctcagccta
     3421 ccgtggtgtt cgtgtctaag aagggcctgc agaagatcct gaacgtgcag aagaagctgc
     3481 ctatcatcca gaagatcatc atcatggact ctaagaccga ctaccagggc ttccagagca
     3541 tgtacacatt cgtgacatct catctgcctc ctggcttcaa cgagtacgac ttcgtgccag
     3601 agtctttcga cagggacaaa accattgccc tgatcatgaa cagctctggg tctaccggcc
     3661 tgcctaaggg cgtggccctg cctcatcgca ccgcctgtgt gcgcttctct cacgcccgcg
     3721 accctatttt cggcaaccag atcatccccg acaccgctat tctgagcgtg gtgccattcc
     3781 accacggctt cggcatgttc accaccctgg gctacctgat ttgcggcttt cgggtggtgc
     3841 tgatgtaccg cttcgaggag gagctgttcc tgcgcagcct gcaagactac aaaattcagt
     3901 ctgccctgct ggtgccaacc ctgttcagct tcttcgctaa gagcaccctg atcgacaagt
     3961 acgacctgtc taacctgcac gagattgcct ctggcggcgc cccactgtct aaggaggtgg
     4021 gcgaagccgt ggccaagcgc tttcatctgc caggcatccg ccagggctac ggcctgaccg
     4081 agacaaccag cgccattctg attaccccag agggcgacga caagcctggc gccgtgggca
     4141 aggtggtgcc attcttcgag gccaaggtgg tggacctgga caccggcaag accctgggag
     4201 tgaaccagcg cggcgagctg tgtgtgcgcg gccctatgat tatgtccggc tacgtgaata
     4261 accctgaggc cacaaacgcc ctgatcgaca aggacggctg gctgcactct ggcgacattg
     4321 cctactggga cgaggacgag cacttcttca tcgtggaccg cctgaagtct ctgatcaagt
     4381 acaagggcta ccaggtggcc ccagccgagc tggagtctat cctgctgcag caccctaaca
     4441 ttttcgacgc cggagtggcc ggcctgcccg acgacgatgc cggcgagctg cctgccgccg
     4501 tcgtcgtgct ggaacacggc aagaccatga ccgagaagga gatcgtggac tatgtggcca
     4561 gccaggtgac aaccgccaag aagctgcgcg gcggagtggt gttcgtggac gaggtgccca
     4621 agggcctgac cggcaagctg gacgcccgca agatccgcga gatcctgatc aaggctaaga
     4681 aaggcggcaa gatcgccgtg taataattct agagtcgggg cggccggccg cttcgagcag
     4741 acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat
     4801 gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata
     4861 aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg
     4921 aggtttttta aagcaagtaa aacctctaca aatgtggtaa aatcgataag gatccaggtg
     4981 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     5041 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     5101 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5161 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5221 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5281 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5341 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5401 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5461 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5521 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     5581 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     5641 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     5701 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     5761 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     5821 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     5881 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     5941 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     6001 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     6061 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     6121 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     6181 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     6241 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     6301 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     6361 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6421 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     6481 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6541 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     6601 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     6661 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     6721 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     6781 acatggctcg ac
//
