LOCUS       sequence-230277-        7407 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 7407)
  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 7407)
  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: hsa-cdk2 3'UTR sicheck2 mut
FEATURES             Location/Qualifiers
     source          1..7407
                     /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    2822..2870
                     /note="synthetic polyadenylation signal"
     promoter        2879..3630
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      3660..3669
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3666..5318
                     /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             4625..4633
                     /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    5362..5483
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5373..5392)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5427..5446
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5616..5720
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5721..5789,5790..6581)
                     /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(5939..5958)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6752..7340
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7241..7260
                     /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 cgctcgagag ccttcttgaa gcccccagcc ctaatctcac
     1681 cctctcctcc agtgtgggct tgaccaggct tggccttggg ctatttggac tcaggtgggc
     1741 cctctgaact tgccttaaac actcaccttc tagtcttggc cagccaactc tgggaataca
     1801 ggggtgaaag gggggaacca gtgaaaatga aaggaagttt cagtattaga tgcacttaag
     1861 ttagcctcca ccaccctttc ccccttctct tagttattgc tgaagagggt tggtataaaa
     1921 ataattttaa aaaagccttc ctacacgtta gatttgccgt accaatctct gaatgcccca
     1981 taattattat ttccagtgtt tgggatgacc aggatcccaa gcctcctgct gccacaatgt
     2041 ttataaaggc caaatgatag cgggggctaa gttggtgctt ttgagaacca agtaaaacaa
     2101 aacgtgaccg aggagtctat tttaaagaat tcggttgaaa aaatagatcc aatcagttta
     2161 taccctagtt agtgttttgc ctcacctaat aggctgggag actgaagact cagcccgggt
     2221 ggggctgcag aaaaatgatt ggccccagtc cccttgtttg tcccttctac aggcatgagg
     2281 aatctgggag gccctgagac agggattgtg cttcattcca atctattgct tcaccatggc
     2341 cttatgaggc aggtgagaga tgtttgaatt tttctcttcc ttttagtatt cttagttgtt
     2401 cagttgccaa ggatccctga tcccattttc ctctgacgtc cacctcctac cccataggag
     2461 ttagaagtta gggtttaggc atcattttga gaatgctgac actttttcag ggctgtgatt
     2521 gagtgagggc atgggtaaaa atatttcttt aaaagaagga tgaacaatta tatttatatt
     2581 tcaggttata tccaatagta gagttggctt tttttttttt tttttggtca tagtgggtgg
     2641 atttgttgcc atgtgcacct tggggttttg taatgacagt gctaaaaaaa aaaagcattt
     2701 tttttttatg atttgtctct gtcacccttg tccttgagtg ctcttgctat taacgttatt
     2761 tgtaatttag tttgtagctc attaaaaaaa tgtgcctagt tttatagcgg ccgctggccg
     2821 caataaaata tctttatttt cattacatct gtgtgttggt tttttgtgtg aggatctaaa
     2881 tgagtcttcg gacctcgcgg gggccgctta agcggtggtt agggtttgtc tgacgcgggg
     2941 ggagggggaa ggaacgaaac actctcattc ggaggcggct cggggtttgg tcttggtggc
     3001 cacgggcacg cagaagagcg ccgcgatcct cttaagcacc cccccgccct ccgtggaggc
     3061 gggggtttgg tcggcgggtg gtaactggcg ggccgctgac tcgggcgggt cgcgcgcccc
     3121 agagtgtgac cttttcggtc tgctcgcaga cccccgggcg gcgccgccgc ggcggcgacg
     3181 ggctcgctgg gtcctaggct ccatggggac cgtatacgtg gacaggctct ggagcatccg
     3241 cacgactgcg gtgatattac cggagacctt ctgcgggacg agccgggtca cgcggctgac
     3301 gcggagcgtc cgttgggcga caaacaccag gacggggcac aggtacacta tcttgtcacc
     3361 cggaggcgcg agggactgca ggagcttcag ggagtggcgc agctgcttca tccccgtggc
     3421 ccgttgctcg cgtttgctgg cggtgtcccc ggaagaaata tatttgcatg tctttagttc
     3481 tatgatgaca caaaccccgc ccagcgtctt gtcattggcg aattcgaaca cgcagatgca
     3541 gtcggggcgg cgcggtccca ggtccacttc gcatattaag gtgacgcgtg tggcctcgaa
     3601 caccgagcga ccctgcagcg acccgcttaa aagcttggca ttccggtact gttggtaaag
     3661 ccaccatggc cgatgctaag aacattaaga agggccctgc tcccttctac cctctggagg
     3721 atggcaccgc tggcgagcag ctgcacaagg ccatgaagag gtatgccctg gtgcctggca
     3781 ccattgcctt caccgatgcc cacattgagg tggacatcac ctatgccgag tacttcgaga
     3841 tgtctgtgcg cctggccgag gccatgaaga ggtacggcct gaacaccaac caccgcatcg
     3901 tggtgtgctc tgagaactct ctgcagttct tcatgccagt gctgggcgcc ctgttcatcg
     3961 gagtggccgt ggcccctgct aacgacattt acaacgagcg cgagctgctg aacagcatgg
     4021 gcatttctca gcctaccgtg gtgttcgtgt ctaagaaggg cctgcagaag atcctgaacg
     4081 tgcagaagaa gctgcctatc atccagaaga tcatcatcat ggactctaag accgactacc
     4141 agggcttcca gagcatgtac acattcgtga catctcatct gcctcctggc ttcaacgagt
     4201 acgacttcgt gccagagtct ttcgacaggg acaaaaccat tgccctgatc atgaacagct
     4261 ctgggtctac cggcctgcct aagggcgtgg ccctgcctca tcgcaccgcc tgtgtgcgct
     4321 tctctcacgc ccgcgaccct attttcggca accagatcat ccccgacacc gctattctga
     4381 gcgtggtgcc attccaccac ggcttcggca tgttcaccac cctgggctac ctgatttgcg
     4441 gctttcgggt ggtgctgatg taccgcttcg aggaggagct gttcctgcgc agcctgcaag
     4501 actacaaaat tcagtctgcc ctgctggtgc caaccctgtt cagcttcttc gctaagagca
     4561 ccctgatcga caagtacgac ctgtctaacc tgcacgagat tgcctctggc ggcgccccac
     4621 tgtctaagga ggtgggcgaa gccgtggcca agcgctttca tctgccaggc atccgccagg
     4681 gctacggcct gaccgagaca accagcgcca ttctgattac cccagagggc gacgacaagc
     4741 ctggcgccgt gggcaaggtg gtgccattct tcgaggccaa ggtggtggac ctggacaccg
     4801 gcaagaccct gggagtgaac cagcgcggcg agctgtgtgt gcgcggccct atgattatgt
     4861 ccggctacgt gaataaccct gaggccacaa acgccctgat cgacaaggac ggctggctgc
     4921 actctggcga cattgcctac tgggacgagg acgagcactt cttcatcgtg gaccgcctga
     4981 agtctctgat caagtacaag ggctaccagg tggccccagc cgagctggag tctatcctgc
     5041 tgcagcaccc taacattttc gacgccggag tggccggcct gcccgacgac gatgccggcg
     5101 agctgcctgc cgccgtcgtc gtgctggaac acggcaagac catgaccgag aaggagatcg
     5161 tggactatgt ggccagccag gtgacaaccg ccaagaagct gcgcggcgga gtggtgttcg
     5221 tggacgaggt gcccaagggc ctgaccggca agctggacgc ccgcaagatc cgcgagatcc
     5281 tgatcaaggc taagaaaggc ggcaagatcg ccgtgtaata attctagagt cggggcggcc
     5341 ggccgcttcg agcagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     5401 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     5461 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     5521 agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtaaaatcg
     5581 ataaggatcc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     5641 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     5701 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     5761 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     5821 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     5881 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     5941 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     6001 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     6061 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     6121 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     6181 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     6241 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     6301 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     6361 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     6421 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     6481 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     6541 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     6601 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     6661 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     6721 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     6781 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     6841 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     6901 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     6961 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     7021 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     7081 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     7141 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     7201 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     7261 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     7321 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     7381 gctggccttt tgctcacatg gctcgac
//
