LOCUS       sequence-422170-        5015 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used for expressing VP16 fused to ZF-1..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5015)
  AUTHORS   Alcantar MA, English MA, Valeri JA, Collins JJ
  TITLE     A high-throughput synthetic biology approach for studying 
            combinatorial chromatin-based transcriptional regulation.
  JOURNAL   Mol Cell. 2024 Jun 20;84(12):2382-2396.e9. doi: 
            10.1016/j.molcel.2024.05.025.
  PUBMED    38906116
REFERENCE   2  (bases 1 to 5015)
  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: 
            "10.1016/j.molcel.2024.05"; journalName: "Mol Cell"; date: 
            "2024-06-20- 20"; volume: "84"; issue: "12"; pages: "2382-2396.e9"
COMMENT     Sequence Label: pPtet-1-VP16
FEATURES             Location/Qualifiers
     source          1..5015
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    232..250
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    271..289
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             313..378
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             385..405
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        1514..1701
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             1702..2364
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     misc_feature    2579..3082
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     CDS             complement(3099..3890)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      4320..4908
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4809..4828
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccctgaattc gcatctagat ggtagagcca caaacagccg gtacaagcaa cgatctccag
       61 gaccatctga atcatgcgcg gatgacacga actcacgacg gcgatcacag acattaaccc
      121 acagtacaga cactgcgaca acgtggcaat tcgtcgcaat accgtctcac tgaactggcc
      181 gataattgca gacgaacgtt tttttctttt tttttttttg cggtataaaa atccctatca
      241 gtgatagaga ttttgttttt tttttttttt tccctatcag tgatagagaa tatcaagcta
      301 caaaaaggta tggattacaa agatcatgat ggtgattata aggaccatga tattgactac
      361 aaagatgacg acgacaaaat ggcccccaag aaaaagagga aggtaggtat ccacggtgtt
      421 ccgggtgggg gttctggttc tagacccggt gaacgtccgt ttcaatgtag aatctgcatg
      481 gcgaactttt ccagacaaga tcgtcttgat cgtcatacca ggacccacac tggcgaaaag
      541 ccctttcaat gtaggatatg tatggcgaac ttcagtcaga aagaacatct ggcggggcat
      601 ctgagaactc atactggtga aaagcccttc cagtgccgta tctgcatggc aaatttttcc
      661 cgtagagata acttaaaccg tcatctgaaa acccatctga gaggttccgg aggttctggc
      721 ggtggaagtg gtggcggagg tagcggtggt ggcagcggtt cctgcagggg ttctctagaa
      781 gctccaccta ctgatgtgtc acttggcgat gagttacacc ttgatggtga agatgtggca
      841 atggcgcatg cggacgccct agacgacttt gatttggaca tgttaggtga cggcgactca
      901 ccaggaccgg gtttcacccc tcacgatagt gccccatacg gtgctttgga catggccgac
      961 tttgagtttg aacaaatgtt tacggatgcc ttaggaattg acgaatacgg tggtggaagc
     1021 agaccaggat cctaactcga gattgaattg aattgaaatc gatagatcaa tttttttctt
     1081 ttctctttcc ccatccttta cgctaaaata atagtttatt ttattttttg aatatttttt
     1141 atttatatac gtatatatag actattattt atcttttaat gattattaag atttttatta
     1201 aaaaaaaatt cgctcctctt ttaatgcctt tatgcagttt ttttttccca ttcgatattt
     1261 ctatgtgctg gaaatctgct cgtcagtggt gctcacactg acgaatcatg tacagatcat
     1321 accgatgact gcctggcgac tcacaactaa gcaagacagc cggaaccagc gccggcgaac
     1381 accactgcat atatggcata tcacaacagt ccacgtctca agcagttaca gagatgttac
     1441 gaaccactag tgcactgcag tacaaacaca gtcctttccc gcaattttct ttttctatta
     1501 ctcttggcct cctctagtac actctatatt tttttatgcc tcggtaatga ttttcatttt
     1561 tttttttcca cctagcggat gactcttttt ttttcttagc gattggcatt atcacataat
     1621 gaattataca ttatataaag taatgtgatt tcttcgaaga atatactaaa aaatgagcag
     1681 gcaagataaa cgaaggcaaa gatgacagag cagaaagccc tagtaaagcg tattacaaat
     1741 gaaaccaaga ttcagattgc gatctcttta aagggtggtc ccctagcgat agagcactcg
     1801 atcttcccag aaaaagaggc agaagcagta gcagaacagg ccacacaatc gcaagtgatt
     1861 aacgtccaca caggtatagg gtttctggac catatgatac atgctctggc caagcattcc
     1921 ggctggtcgc taatcgttga gtgcattggt gacttacaca tagacgacca tcacaccact
     1981 gaggactgcg ggattgctct cggtcaagct tttaaagagg ccctaggggc cgtgcgtgga
     2041 gtaaaaaggt ttggatcagg atttgcgcct ttggatgagg cactttccag agcggtggtt
     2101 gatctttcga acaggccgta cgcagttgtc gaacttggtt tgcaaaggga gaaagtaggt
     2161 gatctctctt gcgagatgat cccgcatttt cttgaaagct ttgcagaggc tagcagaatt
     2221 accctccacg ttgattgtct gcgaggcaag aatgatcatc accgtagtga gagtgcgttc
     2281 aaggctcttg cggttgccat aagagaagcc acctcgccca atggtaccaa cgatgttccc
     2341 tccaccaaag gtgttcttat gtagtgacac cgattattta aagctgctgc atacgatata
     2401 tatacatgtg tatatatgta tacctatgaa tgtcagtaag tatgtatacg aacagtatga
     2461 tactgaagat gacaaggtaa tgcatcattc tatacgtgtc attctgaacg aggcgcgctt
     2521 tccttttttc tttttgcttt ttcttttttt ttctcttgaa ctcgacggat catagagtat
     2581 cacgtgctat aaaaataatt ataatttaaa ttttttaata taaatatata aattaaaaat
     2641 agaaagtaaa aaaagaaatt aaagaaaaaa tagtttttgt tttccgaaga tgtaaaagac
     2701 tctaggggga tcgccaacaa atactacctt ttatcttgct cttcctgctc tcaggtatta
     2761 atgccgaatt gtttcatctt gtctgtgtag aaaaccacac acgaaaatcc tgtgatttta
     2821 cattttactt atcgttaatc gaatgtatat ctatttaatc tgcttttctt gtctaataaa
     2881 tatatatgta aagtacgctt tttgttgaaa ttttttaaac ctttgtttat ttttttttct
     2941 tcattccgta actcttctac cttctttatt tactttctaa aatccaaata caaaacataa
     3001 aaataaataa acacagagta aattcccaaa ttattccatc attaaaagat acgaggcgcg
     3061 tgtaagttac aggcaagcga tccgtcccga gcggccgctt atttgccgac taccttggtg
     3121 atctcgcctt tcacgtagtg gacaaattct tccaactgat ctgcgcgcga ggccaagcga
     3181 tcttcttctt gtccaagata agcctgtcta gcttcaagta tgacgggctg atactgggcc
     3241 ggcaggcgct ccattgccca gtcggcagcg acatccttcg gcgcgatttt gccggttact
     3301 gcgctgtacc aaatgcggga caacgtaagc actacatttc gctcatcgcc agcccagtcg
     3361 ggcggcgagt tccatagcgt taaggtttca tttagcgcct caaatagatc ctgttcagga
     3421 accggatcaa agagttcctc cgccgctgga cctaccaagg caacgctatg ttctcttgct
     3481 tttgtcagca agatagccag atcaatgtcg atcgtggctg gctcgaagat acccgcaaga
     3541 atgtcattgc gctgccattc tccaaattgc agttcgcgct tagctggata acgccacgga
     3601 atgatgtcgt cgtgcacaac aatggtgact tctacagcgc ggagaatctc gctctctcca
     3661 ggggaagccg aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt ttcatcaagc
     3721 cttacggtca ccgtaaccag caaatcaata tcactgtgtg gcttcaggcc gccatccact
     3781 gcggagccgt acaaatgtac ggccagcaac gtcggttcga gatggcgctc gatgacgcca
     3841 actacctctg atagttgagt cgatacttcg gcgatcaccg cttccctcat aatgtttaac
     3901 tttgttttag ggcgactgcc ctgctgcgta acatcgttgc tgctccataa catcaaacat
     3961 cgacccacgg cgtaacgcgc ttgctgcttg gatgcccgag gcatagactg taccccaaaa
     4021 aaacagtcat aacaagccat gaaaaccgcc actgcgccgt taccaccgct gcgttcggtc
     4081 aaggttctgg accagttgcg tgagcgcata cgctacttgc attacagctt acgaaccgaa
     4141 caggcttatg tccactgggt tcgtgccttc atccgtttcc acggtgtgcg tcacccggca
     4201 accttgggta gcagcgaagt cgaggcattt ctgtcctggc tggtcatgac caaaatccct
     4261 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4321 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4381 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4441 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     4501 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4561 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4621 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4681 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4741 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4801 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4861 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4921 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     4981 ttatcccctg attctgtgga taaccgtgcg gccgc
//
