LOCUS       sequence-426757-        5014 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used for expressing VP16 fused to ZF-2..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5014)
  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 5014)
  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-2-VP16
FEATURES             Location/Qualifiers
     source          1..5014
                     /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        1513..1700
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             1701..2363
                     /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    2578..3081
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     CDS             complement(3098..3889)
                     /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      4319..4907
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4808..4827
                     /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 tttttatttt ttgttttttg cggtataaaa atccctatca
      241 gtgatagaga ttttattttt tttttctttt tccctatcag tgatagagaa tatcaagcta
      301 caaaaaggta tggattataa agaccatgac ggagattata aagatcatga catagattac
      361 aaggatgacg atgacaagat ggcaccaaag aagaagagga aggtgggtat tcacggagtc
      421 cccggaggtg gctcagggtc ccgtcccggt gagaggccct ttcaatgcag aatttgcatg
      481 gcaaactttt ccacggggca aatcttggat cgtcatacaa gaacgcatac gggcgaaaaa
      541 ccattccaat gccgtatttg catggctaac tttagcgttg cgcactcctt gaagaggcac
      601 ctgagaacac atactggcga gaagcccttt cagtgcagaa tctgtatggc gaatttctcc
      661 gatccctcca atcttcgtag acatttgaaa actcatttaa gaggatctgg cggtagtggc
      721 ggagggagtg gtggcggggg ctctggaggc ggatctgggt 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 gataaagcaa tcttgatgag gataatgatt tttttttgaa
     1081 tatacataaa tactaccgtt tttctgctag attttgtgat gacgtaaata agtacatatt
     1141 actttttaag ccaagacaag attaagcatt aactttaccc ttttctttct aagtttcaat
     1201 attagttatc actgtttaaa agttatggcg agaacgtcgg cggttaaaat atattaccct
     1261 gaacggctgg aaatctgctc gtcagtggtg ctcacactga cgaatcatgt acagatcata
     1321 ccgatgactg cctggcgact cacaactaag caagacagcc ggaaccagcg ccggcgaaca
     1381 ccactgcata tatggcatat cacaacagtc cacgtctcaa gcagttacag agatgttacg
     1441 aaccactagt gcactgcagt acaaacacag tcctttcccg caattttctt tttctattac
     1501 tcttggcctc ctctagtaca ctctatattt ttttatgcct cggtaatgat tttcattttt
     1561 ttttttccac ctagcggatg actctttttt tttcttagcg attggcatta tcacataatg
     1621 aattatacat tatataaagt aatgtgattt cttcgaagaa tatactaaaa aatgagcagg
     1681 caagataaac gaaggcaaag atgacagagc agaaagccct agtaaagcgt attacaaatg
     1741 aaaccaagat tcagattgcg atctctttaa agggtggtcc cctagcgata gagcactcga
     1801 tcttcccaga aaaagaggca gaagcagtag cagaacaggc cacacaatcg caagtgatta
     1861 acgtccacac aggtataggg tttctggacc atatgataca tgctctggcc aagcattccg
     1921 gctggtcgct aatcgttgag tgcattggtg acttacacat agacgaccat cacaccactg
     1981 aggactgcgg gattgctctc ggtcaagctt ttaaagaggc cctaggggcc gtgcgtggag
     2041 taaaaaggtt tggatcagga tttgcgcctt tggatgaggc actttccaga gcggtggttg
     2101 atctttcgaa caggccgtac gcagttgtcg aacttggttt gcaaagggag aaagtaggtg
     2161 atctctcttg cgagatgatc ccgcattttc ttgaaagctt tgcagaggct agcagaatta
     2221 ccctccacgt tgattgtctg cgaggcaaga atgatcatca ccgtagtgag agtgcgttca
     2281 aggctcttgc ggttgccata agagaagcca cctcgcccaa tggtaccaac gatgttccct
     2341 ccaccaaagg tgttcttatg tagtgacacc gattatttaa agctgctgca tacgatatat
     2401 atacatgtgt atatatgtat acctatgaat gtcagtaagt atgtatacga acagtatgat
     2461 actgaagatg acaaggtaat gcatcattct atacgtgtca ttctgaacga ggcgcgcttt
     2521 ccttttttct ttttgctttt tctttttttt tctcttgaac tcgacggatc atagagtatc
     2581 acgtgctata aaaataatta taatttaaat tttttaatat aaatatataa attaaaaata
     2641 gaaagtaaaa aaagaaatta aagaaaaaat agtttttgtt ttccgaagat gtaaaagact
     2701 ctagggggat cgccaacaaa tactaccttt tatcttgctc ttcctgctct caggtattaa
     2761 tgccgaattg tttcatcttg tctgtgtaga aaaccacaca cgaaaatcct gtgattttac
     2821 attttactta tcgttaatcg aatgtatatc tatttaatct gcttttcttg tctaataaat
     2881 atatatgtaa agtacgcttt ttgttgaaat tttttaaacc tttgtttatt tttttttctt
     2941 cattccgtaa ctcttctacc ttctttattt actttctaaa atccaaatac aaaacataaa
     3001 aataaataaa cacagagtaa attcccaaat tattccatca ttaaaagata cgaggcgcgt
     3061 gtaagttaca ggcaagcgat ccgtcccgag cggccgctta tttgccgact accttggtga
     3121 tctcgccttt cacgtagtgg acaaattctt ccaactgatc tgcgcgcgag gccaagcgat
     3181 cttcttcttg tccaagataa gcctgtctag cttcaagtat gacgggctga tactgggccg
     3241 gcaggcgctc cattgcccag tcggcagcga catccttcgg cgcgattttg ccggttactg
     3301 cgctgtacca aatgcgggac aacgtaagca ctacatttcg ctcatcgcca gcccagtcgg
     3361 gcggcgagtt ccatagcgtt aaggtttcat ttagcgcctc aaatagatcc tgttcaggaa
     3421 ccggatcaaa gagttcctcc gccgctggac ctaccaaggc aacgctatgt tctcttgctt
     3481 ttgtcagcaa gatagccaga tcaatgtcga tcgtggctgg ctcgaagata cccgcaagaa
     3541 tgtcattgcg ctgccattct ccaaattgca gttcgcgctt agctggataa cgccacggaa
     3601 tgatgtcgtc gtgcacaaca atggtgactt ctacagcgcg gagaatctcg ctctctccag
     3661 gggaagccga agtttccaaa aggtcgttga tcaaagctcg ccgcgttgtt tcatcaagcc
     3721 ttacggtcac cgtaaccagc aaatcaatat cactgtgtgg cttcaggccg ccatccactg
     3781 cggagccgta caaatgtacg gccagcaacg tcggttcgag atggcgctcg atgacgccaa
     3841 ctacctctga tagttgagtc gatacttcgg cgatcaccgc ttccctcata atgtttaact
     3901 ttgttttagg gcgactgccc tgctgcgtaa catcgttgct gctccataac atcaaacatc
     3961 gacccacggc gtaacgcgct tgctgcttgg atgcccgagg catagactgt accccaaaaa
     4021 aacagtcata acaagccatg aaaaccgcca ctgcgccgtt accaccgctg cgttcggtca
     4081 aggttctgga ccagttgcgt gagcgcatac gctacttgca ttacagctta cgaaccgaac
     4141 aggcttatgt ccactgggtt cgtgccttca tccgtttcca cggtgtgcgt cacccggcaa
     4201 ccttgggtag cagcgaagtc gaggcatttc tgtcctggct ggtcatgacc aaaatccctt
     4261 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     4321 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     4381 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     4441 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     4501 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     4561 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     4621 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     4681 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     4741 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     4801 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     4861 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     4921 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     4981 tatcccctga ttctgtggat aaccgtgcgg ccgc
//
