LOCUS       sequence-252230-        9369 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Repressor plasmid for WTC 846. Encodes TetR and TetR-Tup1, His3MX 
            selection marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9369)
  AUTHORS   Azizoglu A, Brent R, Rudolf F
  TITLE     A precisely adjustable, variation-suppressed eukaryotic 
            transcriptional controller to enable genetic discovery.
  JOURNAL   Elife. 2021 Aug 3;10. pii: 69549. doi: 10.7554/eLife.69549.
  PUBMED    34342575
REFERENCE   2  (bases 1 to 9369)
  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: "Elife."; 
            date: "2021-08-3"; volume: "10. pii: 69549. doi"; pages: " 
            10.7554/eLife.69549"
COMMENT     Sequence Label: FRP2371
FEATURES             Location/Qualifiers
     source          1..9369
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     24..43
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        24..42
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(954..973)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     CDS             1499..1519
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      3831..4080
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     CDS             5514..5534
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      5577..5766
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     promoter        complement(5852..5870)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5891..5907)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5891..5907)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5904..5926)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5915..5931
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     complement(6165..6182)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6336..6924)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6416..6435)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7103..7894,7895..7963))
                     /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     7726..7745
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7964..8068)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             8366..9016
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his5"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=S. pombe his5
                     /note="yeast auxotrophic marker; corresponds to S. 
                     cerevisiae HIS3"
                     /translation="MRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHANQK
                     GEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQA
                     MGNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLYS
                     FSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
     primer_bind     join(9355..9369,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(9369,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgaa ttgtaatacg actcactata gggcgaattg gagaaacaga
       61 ataatatata aacctgtata atataacctt gaagactata tttcttttcg cggccgccta
      121 tagggcgaat tggagctcca ccgcggtggc tctagaagtc gaacaagaag caggcaaagt
      181 ttagagcact gcccctccgc actcaaaaaa gaaaaaacta ggaggaaaat aaaattctca
      241 accacacaaa cacataaaca catacaaata caaatacaag cttatttact tgacatcgcg
      301 cgatcttcca ctattcagcg ccgtccgccc tctctcgtgt tttttgttta cgcgacaact
      361 atgcgaaatc cggagcaacg ggcaaccgtt tggggaaaga ccacacccac gcgcgatcgc
      421 catggcaacg aggtcgcaca cgccccacac ccagacctcc ctgcgagcgg gcatgggtac
      481 aatgtccccg ttgccacaga gaccacttcg tagcacagcg cagagcgtag cgtgttgttg
      541 ctgctgacaa aagaaaattt ttcttagcaa agcaaaggag gggaagcacg ggcagatagc
      601 accgtaccat acccttggaa actcgaaatg aacgaagcag gaaatgagag aatgagagtt
      661 ttgtaggtat atatagcggt agtgtttgcg cgttaccatc atcttctgga tctatctatt
      721 gttcttttcc tcatcacttt cccctttttc gctcttcttc ttgtctttta tttctttctt
      781 ttttttaatt gttccctcga ttggctatct accaaagaat ccaaacttaa tacacgtatt
      841 tatttgtcca attaccggat cccccgggct gcaggaattc atgtccagat tagataaaag
      901 taaagtgatt aacagcgcat tagagctgct taatgaggtc ggaatcgaag gtttaacaac
      961 ccgtaaactc gcccagaagc taggtgtaga gcagcctaca ttgtattggc atgtaaaaaa
     1021 taagcgggct ttgctcgacg ccttagccat tgagatgtta gataggcacc atactcactt
     1081 ttgcccttta gaaggggaaa gctggcaaga ttttttacgt aataacgcta aaagttttag
     1141 atgtgcttta ctaagtcatc gcgatggagc aaaagtacat ttaggtacac ggcctacaga
     1201 aaaacagtat gaaactctcg aaaatcaatt agccttttta tgccaacaag gtttttcact
     1261 agagaatgca ttatatgcac tcagcgctgt ggggcatttt actttaggtt gcgtattgga
     1321 agatcaagag catcaagtcg ctaaagaaga aagggaaaca cctactactg atagtatgcc
     1381 gccattatta cgacaagcta tcgaattatt tgatcaccaa ggtgcagagc cagccttctt
     1441 attcggcctt gaattgatca tttgcggatt agaaaaacaa cttaaatgtg aaagtgcccc
     1501 caagaaaaag agaaaggtgg cagaagccgc cgcaaaggaa gcggcagcaa aagaggctgc
     1561 ggccaaggaa gctgctgcca aggcgctaga agcagaggcg gccgcaaagg aggcggccgc
     1621 gaaggaagcg gctgcgaagg aggcagcggc aaaggcgatg actgccagcg tttcgaatac
     1681 gcagaataag ctgaatgagc ttctcgatgc catcagacag gagtttctcc aagtctcaca
     1741 agaggcaaat acctaccgtc ttcaaaacca aaaggattac gatttcaaaa tgaaccagca
     1801 gctggctgag atgcagcaga taagaaacac cgtctacgaa ctggaactaa ctcacaggaa
     1861 aatgaaggac gcgtacgaag aagagatcaa gcacttgaaa ctagggctgg agcaaagaga
     1921 ccatcaaatt gcatctttga ccgtccagca acagcggcaa cagcaacagc agcaacaggt
     1981 ccagcagcat ttacaacagc aacagcagca gctagccgct gcatctgcat ctgttccagt
     2041 tgcgcaacaa ccaccggcta ctacttcggc caccgccact ccagcagcaa acacaactac
     2101 tggttcgcca tcggccttcc cagtacaagc tagccgtcct aatctggttg gctcacagtt
     2161 gcctaccacc actttgcctg tggtgtcctc aaacgcccaa caacaactac cacaacagca
     2221 actgcaacag cagcaacttc aacaacagca accacctccc caggtttccg tggcaccatt
     2281 gagtaacaca gccatcaacg gatctcctac ttctaaagag accactactt taccctctgt
     2341 caaggcacct gaatctacgt tgaaagaaac tgaaccggaa aataataata cctcgaagat
     2401 aaatgacacc ggatccgcca ccacggccac cactaccacc gcaactgaaa ctgaaatcaa
     2461 acctaaggag gaagacgcca ccccggctag tttgcaccag gatcactact tagtccctta
     2521 taatcaaaga gcaaaccact ctaaacctat cccacctttc cttttggatc tagattccca
     2581 gtctgttccc gatgctctga agaagcaaac aaatgattat tatattttat acaacccggc
     2641 actaccaaga gaaattgacg ttgagttaca caaatctttg gatcatactt cagttgtttg
     2701 ttgcgtgaag ttcagtaacg atggtgaata cttagccaca ggctgcaaca aaactactca
     2761 agtgtatcgc gtttcagatg gttctctggt ggcccgtcta tctgacgatt ctgctgccaa
     2821 taaccatcga aattcgatca ctgaaaataa caccaccacg tccacggata acaatacaat
     2881 gacaaccact actaccacca caattactac cacagcgatg acttcggcag cagaattggc
     2941 aaaagatgtg gaaaacctga acacttcgtc ttccccatca tccgacttgt atatccgttc
     3001 agtgtgtttt tctccagatg ggaaattttt ggcaacaggt gctgaagaca gactgattag
     3061 aatttgggat attgaaaata gaaagattgt tatgattctt caaggccacg aacaagatat
     3121 ttattcattg gactactttc cctcaggtga caaattagtc tccggttctg gtgaccgtac
     3181 cgttcgtatt tgggacttac gtacaggcca gtgttcattg actttatcca ttgaagatgg
     3241 tgttaccacc gtcgctgtat caccaggtga tggtaaatac atcgctgctg gttctctaga
     3301 tcgtgctgtg agagtttggg attccgagac cggattcttg gtggaaagac tagattcgga
     3361 aaacgaatcc ggtacaggcc acaaggactc tgtttatagc gttgtcttca ctagagatgg
     3421 acaaagcgtt gtatccggct cattagatag atctgttaag ctctggaatt tgcagaatgc
     3481 aaacaacaag agcgattcga aaactccaaa ttccggcact tgtgaagtta cgtatatcgg
     3541 gcataaagac tttgtattgt ccgtggccac cacacaaaat gatgagtaca tcttgtccgg
     3601 ttccaaagat cgtggtgtcc tgttttggga taagaaatcc ggcaatccgt tattgatgtt
     3661 gcaaggtcat aggaattcag ttatatctgt ggctgtggca aacgggtctc cgctgggtcc
     3721 agaatataac gtttttgcta ctggtagcgg tgattgtaaa gcaaggattt ggaagtataa
     3781 aaaaatagcg ccaaattaac tcgagacagg ccccttttcc tttgtctata tcatgtaatt
     3841 agttatgtca cgcttacatt cacgccctcc tcccacatcc gctctaaccg aaaaggaagg
     3901 agttagacaa cctgaagtct aggtccctat ttattttttt taatagttat gttagtatta
     3961 agaacgttat ttatatttca aatttttctt ttttttctgt acaaacgcgt gtacgcatgt
     4021 aacattatac tgaaaacctt gcttgagaag gttttgggac gctcgaaggc tttaatttgc
     4081 aaggttcgca gtttacactc tcatcgtcgc tctcatcatc gcttccgttg ttgttttcct
     4141 tagtagcgtc tgctttctag acagttcgag tttatcatta tcaatactgc catttcaaag
     4201 aatacgtaaa taattaatag tagtgatttt cctaacttta tttagtcaaa aaattagcct
     4261 ctatcattga tagagtgtct gggtgatcta tcattgatag agcatccact aattttaatt
     4321 ctgctgtaac ccgtacatgc ccaaaatagg gggcgggtta cacagaatct atcattgata
     4381 gagtgtctgg gtgatctatc attgatagag catccactaa atataatgga gctctatcat
     4441 tgatagagca tccactaaaa aaaagaatcc cagcaccaaa atattgtttt cttcaccaac
     4501 catcagttca taggtccatt ctcttagcgc aactacagag aacaggggca caaacaggca
     4561 aaaaacgggc acaacctcaa tggagtgatg caacctgcct ggagtaaatg atgacacaag
     4621 gcaattgacc cacgcatgta tctatctcat tttcttacac cttctattac cttctgctct
     4681 ctctgatttg gaaaaagctg aaaaaaaagg ttgaaaccag ttccctgaaa ttattcccct
     4741 atctatcatt gatagatata aatatctatc attgatagag taattctgta aatctatttc
     4801 ttaaacttct taaattctac ttttatagtt agtctttttt ttagttttaa aacaccaaga
     4861 acttagtttc gaataaacac acataaacaa acaaaatgtc caggttggac aagtccaagg
     4921 ttattaactc tgccttggag ttgttgaacg aagttggtat tgaaggtttg accactagaa
     4981 agttggctca aaaattgggt gttgaacaac ctaccttgta ctggcatgtt aagaacaaaa
     5041 gagctttgtt ggacgctttg gccattgaaa tgttggatag acatcatacc catttctgtc
     5101 cattggaagg tgaatcatgg caagatttct tgagaaacaa cgccaagtct ttcagatgtg
     5161 ctttgttgtc tcatagagat ggtgctaaag ttcacttggg tactagacca actgaaaagc
     5221 aatacgaaac tttggaaaac cagttggctt tcttgtgtca acaaggtttc tctttagaaa
     5281 acgccttgta tgctttgtct gctgttggtc attttacctt gggttgtgtt ttggaagatc
     5341 aagaacatca agtcgccaaa gaagaaaggg aaactccaac tactgattct atgccaccat
     5401 tattgagaca agccattgag ttgtttgatc atcaaggtgc tgaaccagct tttttgtttg
     5461 gtttggaatt gatcatctgc ggtttggaga aacaattgaa gtgtgaatct gcccctaaga
     5521 agaagagaaa ggtttgaatt agttatgtca cgcttacatt cacgccctcc ccccacatcc
     5581 gctctaaccg aaaaggaagg agttagacaa cctgaagtct aggtccctat ttattttttt
     5641 atagttatgt tagtattaag aacgttattt atatttcaaa tttttctttt ttttctgtac
     5701 agacgcgtgt acgcatgtaa cattatactg aaaaccttgc ttgagaaggt tttgggacgc
     5761 tcgaaggctt taatttgggc cgtatttaca gataacgatg tcgttattat atatatatat
     5821 atatattgtc acttgtaccc agcttttgtt ccctttagtg agggttaatt tcgagcttgg
     5881 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca acagctgaca
     5941 ccgattattt aaagctgcag catacgatat atatacatgt gtatatatgt atacctatga
     6001 atgtcagtaa gtatgtatac gaacagtatg atactgaaga tgacaaggta atgcatcatt
     6061 ctatacgtgt cattctgaac gaggcgcgcc tctccctgca ggcattaatg aatcggccaa
     6121 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     6181 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     6241 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     6301 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     6361 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     6421 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     6481 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     6541 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6601 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6661 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6721 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     6781 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6841 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6901 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttgtttaaa cctacggggt
     6961 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     7021 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     7081 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     7141 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     7201 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     7261 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     7321 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     7381 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     7441 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     7501 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatggtt gtcagaagta
     7561 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     7621 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     7681 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     7741 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     7801 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     7861 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     7921 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     7981 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     8041 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     8101 aaagaacgcg gcattagcaa catttgcgaa gcgaacgatt ttgcgaactg ctattaatta
     8161 actctggcgc gcctcctcta gtacactcta tattttttta tgcctcggta atgattttca
     8221 tttttttttt tccacctagc ggatgactct ttttttttct tagcgattgg cattatcaca
     8281 taatgaatta tacattatat aaagtaatgt gatttcttcg aagaatatac taaaaaatga
     8341 gcaggcaaga taaacgaagg cacatatgag gagggctttt gtagaaagaa atacgaacga
     8401 aacgaaaatc agcgttgcca tcgctttgga caaagctccc ttacctgaag agtcgaattt
     8461 tattgatgaa cttataactt ccaagcatgc aaaccaaaag ggagaacaag taatccaagt
     8521 agacacggga attggattct tggatcacat gtatcatgca ctggctaaac atgcaggctg
     8581 gagcttacga ctttactcaa gaggtgattt aatcatcgat gatcatcaca ctgcagaaga
     8641 tactgctatt gcacttggta ttgcattcaa gcaggctatg ggtaactttg ccggtgttaa
     8701 aagatttgga catgcttatt gtccacttga cgaagctctt tccagaagcg tagttgactt
     8761 gtcgggacgg ccctatgctg ttatcgattt gggattaaag cgtgaaaagg ttggggaatt
     8821 gtcctgtgaa atgatccctc acttactata ttccttttcg gtagctgctg gaattacttt
     8881 gcatgttacc tgcttatatg gtagtaatga ccatcatcgt gctgaaagcg cttttaaatc
     8941 tctggctgtt gccatgcgcg cggctacttc tcttactgga agttctgaag tcccaagcac
     9001 gaagggagtg ttgtaaagat gaattggatt atgtcaggaa aagaacgaca attttgcatc
     9061 caaattgtct aaattttaga gttgcttgaa aacaatagaa ccttacttgc tttataatta
     9121 cgttaattag aagcgttatc tcgtgaagga atatagtacg tagccgtata aattgaattg
     9181 aatgttcagc ttatagaata gagacacttt gctgttcaat gcgtcgtcac ttaccatact
     9241 cactttatta tacgacttta agtataaact ccgcggttat ggtaaaatta atgatgcaca
     9301 aacgtccgat cggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
     9361 cacgacgtt
//
