LOCUS       sequence-467804-        5556 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Measure the activity of the reduced differentially methylated region
            (rDMR) of Ctgf with luciferase assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5556)
  AUTHORS   Wu BK, Mei SC, Chen EH, Zheng Y, Pan D
  TITLE     YAP induces an oncogenic transcriptional program through 
            TET1-mediated epigenetic remodeling in liver growth and 
            tumorigenesis.
  JOURNAL   Nat Genet. 2022 Aug;54(8):1202-1213. doi: 
            10.1038/s41588-022-01119-7. Epub 2022 Jul 14.
  PUBMED    35835915
REFERENCE   2  (bases 1 to 5556)
  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"; doi: 
            "10.1038/s41588-022-01119-7"; journalName: "Nat Genet"; date: 
            "2022-08"; volume: "54"; issue: "8"; pages: "1202-1213"
COMMENT     Sequence Label: pGL3-Ctgf-rDMR
FEATURES             Location/Qualifiers
     source          1..5556
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        594..790
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      641..776
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      820..829
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             826..2478
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(892..910)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    2519..2640
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2530..2549)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2584..2603
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2888..2905)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3059..3647)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3139..3158)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3818..4609,4610..4678))
                     /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     4441..4460
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4679..4783)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4810..5265
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4897..4916)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5107..5128
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    5396..5444
                     /note="synthetic polyadenylation signal"
     misc_feature    5458..5549
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     5498..5517
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggtaccgagc tcttacgcgt agcagcatcc atgaggaacc caggtcagcc aattgtatgt
       61 tggtcacttc cccatggtct ctcaccatgc actcagctct ctcctctcag gtaccacatg
      121 cagacacccc cactgacaga cagtgagtgc atagtccatg catgggagca ctcagcgctc
      181 catgttcagg ctcacgaccc attctacagc attggtaaaa ttcattcaga gctttcagcg
      241 ttctatggct ccccagctca tacaactatg aacgtcattt atttattctc agaacacatt
      301 tctggcttct gcaaacacat cagagtcttc ttgcaagcaa gctccaggtg actgctcata
      361 ttccaccata tgttaacaca acactagaaa taaactgccc aaaacactgt gaattatctt
      421 tgttattcag aggtacacaa gaataaattt ctagctttcc atttggttct ttggaatggg
      481 tttagcaatc tgagagaatt tcacactgaa atattcttac acctaaacct tgaggaccct
      541 gacaattatt tttaaaaatt ccatcagccc gtcagtcctc gagatctgcg atctgcatct
      601 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
      661 cagttccgcc cattctccgc cccatcgctg actaattttt tttatttatg cagaggccga
      721 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
      781 cttttgcaaa aagcttggca ttccggtact gttggtaaag ccaccatgga agacgccaaa
      841 aacataaaga aaggcccggc gccattctat ccgctggaag atggaaccgc tggagagcaa
      901 ctgcataagg ctatgaagag atacgccctg gttcctggaa caattgcttt tacagatgca
      961 catatcgagg tggacatcac ttacgctgag tacttcgaaa tgtccgttcg gttggcagaa
     1021 gctatgaaac gatatgggct gaatacaaat cacagaatcg tcgtatgcag tgaaaactct
     1081 cttcaattct ttatgccggt gttgggcgcg ttatttatcg gagttgcagt tgcgcccgcg
     1141 aacgacattt ataatgaacg tgaattgctc aacagtatgg gcatttcgca gcctaccgtg
     1201 gtgttcgttt ccaaaaaggg gttgcaaaaa attttgaacg tgcaaaaaaa gctcccaatc
     1261 atccaaaaaa ttattatcat ggattctaaa acggattacc agggatttca gtcgatgtac
     1321 acgttcgtca catctcatct acctcccggt tttaatgaat acgattttgt gccagagtcc
     1381 ttcgataggg acaagacaat tgcactgatc atgaactcct ctggatctac tggtctgcct
     1441 aaaggtgtcg ctctgcctca tagaactgcc tgcgtgagat tctcgcatgc cagagatcct
     1501 atttttggca atcaaatcat tccggatact gcgattttaa gtgttgttcc attccatcac
     1561 ggttttggaa tgtttactac actcggatat ttgatatgtg gatttcgagt cgtcttaatg
     1621 tatagatttg aagaagagct gtttctgagg agccttcagg attacaagat tcaaagtgcg
     1681 ctgctggtgc caaccctatt ctccttcttc gccaaaagca ctctgattga caaatacgat
     1741 ttatctaatt tacacgaaat tgcttctggt ggcgctcccc tctctaagga agtcggggaa
     1801 gcggttgcca agaggttcca tctgccaggt atcaggcaag gatatgggct cactgagact
     1861 acatcagcta ttctgattac acccgagggg gatgataaac cgggcgcggt cggtaaagtt
     1921 gttccatttt ttgaagcgaa ggttgtggat ctggataccg ggaaaacgct gggcgttaat
     1981 caaagaggcg aactgtgtgt gagaggtcct atgattatgt ccggttatgt aaacaatccg
     2041 gaagcgacca acgccttgat tgacaaggat ggatggctac attctggaga catagcttac
     2101 tgggacgaag acgaacactt cttcatcgtt gaccgcctga agtctctgat taagtacaaa
     2161 ggctatcagg tggctcccgc tgaattggaa tccatcttgc tccaacaccc caacatcttc
     2221 gacgcaggtg tcgcaggtct tcccgacgat gacgccggtg aacttcccgc cgccgttgtt
     2281 gttttggagc acggaaagac gatgacggaa aaagagatcg tggattacgt cgccagtcaa
     2341 gtaacaaccg cgaaaaagtt gcgcggagga gttgtgtttg tggacgaagt accgaaaggt
     2401 cttaccggaa aactcgacgc aagaaaaatc agagagatcc tcataaaggc caagaagggc
     2461 ggaaagatcg ccgtgtaatt ctagagtcgg ggcggccggc cgcttcgagc agacatgata
     2521 agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt
     2581 tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt
     2641 aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt
     2701 taaagcaagt aaaacctcta caaatgtggt aaaatcgata aggatccgtc gaccgatgcc
     2761 cttgagagcc ttcaacccag tcagctcctt ccggtgggcg cggggcatga ctatcgtcgc
     2821 cgcacttatg actgtcttct ttatcatgca actcgtagga caggtgccgg cagcgctctt
     2881 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     2941 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     3001 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     3061 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     3121 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     3181 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     3241 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     3301 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     3361 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     3421 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     3481 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     3541 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     3601 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     3661 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     3721 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     3781 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     3841 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     3901 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     3961 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     4021 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     4081 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     4141 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     4201 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     4261 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     4321 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     4381 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     4441 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     4501 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     4561 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     4621 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     4681 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     4741 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     4801 tgccacctga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     4861 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     4921 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     4981 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     5041 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     5101 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     5161 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     5221 aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttgccat tcgccattca
     5281 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagccca
     5341 agctaccatg ataagtaagt aatattaagg tacgggaggt acttggagcg gccgcaataa
     5401 aatatcttta ttttcattac atctgtgtgt tggttttttg tgtgaatcga tagtactaac
     5461 atacgctctc catcaaaaca aaacgaaaca aaacaaacta gcaaaatagg ctgtccccag
     5521 tgcaagtgca ggtgccagaa catttctcta tcgata
//
