LOCUS       sequence-250430-        7596 bp    DNA     circular SYN 14-APR-2025
DEFINITION  A nuclear plasmid encoding the WT TP-DNAP1 for OrthoRep.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7596)
  AUTHORS   Ravikumar A, Arzumanyan GA, Obadi MKA, Javanpour AA, Liu CC
  TITLE     Scalable, Continuous Evolution of Genes at Mutation Rates above 
            Genomic Error Thresholds.
  JOURNAL   Cell. 2018 Dec 13;175(7):1946-1957.e13. doi: 
            10.1016/j.cell.2018.10.021. Epub 2018 Nov 8.
  PUBMED    30415839
REFERENCE   2  (bases 1 to 7596)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cell.2018.10.021"; journalName: "Cell"; date: 
            "2018-12-13- 13"; volume: "175"; issue: "7"; pages: "1946-1957.e13"
COMMENT     Sequence Label: pAR-Ec318
FEATURES             Location/Qualifiers
     source          1..7596
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      3677..3842
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(3732..3756)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     rep_origin      complement(3885..4473)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4687..4706)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4698..5513
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             4698..5513
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4771..4790)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_feature    5551..6054
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     6096..6114
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6152..6174)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6274..6293
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        6478..6665
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             6666..7328
                     /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"
ORIGIN
        1 gtgttgttat ccgatacaac cggatatttt tcttttaatg agtctaaacc gtgatagctt
       61 caggttaata caatcaaaaa aagctcaaat attcttttaa tgccgcgttc acagattcca
      121 attgaataca actaggtagt tcattatatg aagcctttgc tactattttt cactatagtc
      181 tgccttcacc ttaatgcaga catccacata ttttaatcac tttaaaataa aaaggaagat
      241 atattagaag ctatgatcca atctgtaagc cagattaaaa ttcacgaact cttctttcat
      301 ttgaattgaa tgctttgagt tggggtagat tatcgcaaat tactcatcac atttattgac
      361 tacgaacttg ctgatgtcct ttttttattt atatttttct tcagtgaagc gatttttttt
      421 ttacacagac caagacggaa aaaagtagct aaggaagaaa acaaaatcat gaaaaaaatg
      481 tgaagtgatc atgcacatcg catcaactta aacattggct tagagatata tagagttaga
      541 gtttacggca acctttaagc accaatacct tttggcatag tctaaagacc tggttcttaa
      601 ttttaaacaa atttaactaa agatttccct atcaaagaag taacgagttg acagattttc
      661 tcaaaataaa tcgatactgc atttctaggc atatccagcg agatctatgg actacaaaga
      721 caaagcatta aacgacttga gaaacgtata cgcagacttc gactccttgc cattggactt
      781 cagacaaatc ttaattaagg atagagctac attgttgcaa aaggaagacg ttgaaaagaa
      841 aattttggaa agacaagaag atgccaaaaa gtacgctgaa tacttgaagc aatcagaaat
      901 cccagaaaga atctccttac ctaacatcaa gagacataag ggtgtatcaa tctcctttga
      961 agaaacatct gaagatatgg tcttggaacc aagacctttt atcttcgacg gtttgaacat
     1021 cagatgtttc agaagagaaa ccatattcag tttgaaaaat aagattttga acatggttaa
     1081 ggaatcttca tcctttaaaa acgtttcaag acaaagtgta tcttttatgt acttcaaaat
     1141 ttttaataag ggtaaagtta ttgcaagtac aaagtctgtt aacatctatg aagataagat
     1201 cgacgaaaga ttggaagatt tgtgtaacaa cttcgatgac gtattgaaaa agattataga
     1261 cgtcacctat ggttacgaat ccttattcgt tagtgaaact tactcttacg taatattcta
     1321 tgctaaatca atctactttc cacaacctag atgcgttaac aactggggta acaacatccc
     1381 aaacatcttg acattcgatt cctttaagct tttcaccgca aacaaaaaca acgttagttg
     1441 tatcaagcaa tgttcaagat ttttgtggca aaaagatttc aacactttag aagaaatgat
     1501 agaatacaag aacggtaata tttgtatcgt cacaccacaa ttgcatatca acgatgttag
     1561 agacatcaag tcattcaacg atatcagatt gtactcagaa tcccctatta aaacattttc
     1621 cgttattgat aacactatca catatttgtt ttacttcaag gaacatttgg gtgttatttt
     1681 taacatcact aagtcaagac atgatagaag agtaacaaaa ttttcaccat tgtccaagtt
     1741 cagtgacgtc aaaaatatca ctgtttgttt cgatatcgaa agttacttcg acccagaaaa
     1801 ggaatctaac caagttaaca tccctttcat ttgttgcgct tcaatcatct ataataaggt
     1861 tattggtaac atcgtcgatt ttgaaggtag agactgtgtc gcacaaatga tcgaatacgt
     1921 tgtagatatt tgcggtgaat tgaacataag ttctgttgaa ttgatcgccc ataacggtgg
     1981 tggttatgat ttccactaca tcttgtcatc catgtataat ccagctgcaa ttaaaaacat
     2041 attgatcaga aacaactctt ttatttcttt taatttcgca catgatggtg tcaagttctc
     2101 tgttaaggac agttactcat ttttgttgtg tagtttggcc aacgcttcta aagcattttt
     2161 aaacgaagaa acttttaaga aaactgattt ccctcatcac gacttgaaaa ctgccgatga
     2221 cttatataag gtctacaagg aatggagttc tgttaacaca gaaattaacc atgtcgttga
     2281 aaaggaaaag ttgttgatca cctctgaaca catcgttaac ttcaccaaga acgataagag
     2341 taagactttg atcgaatggt ctaaggatta ctgtagaaac gacgtattgg tcttgtctaa
     2401 ggtttggttg gagtttaaaa acgctgtaga agatattttt aactgcgaat tggtagacca
     2461 aaccatgact ttggctggtt tgtcatacaa gttgttccaa gcaaacatgc cattcgatgt
     2521 cgaattgaga catcctaata aggaagacta cttcaacatg agagaagcat tgattggtgg
     2581 tagatgtatc tctgttaacg gtatctacaa ggatgttttg tgcttagacg taaaatcatt
     2641 atatccagct tctatggctt tctatgatca accttacggt tcttttaaga gagtttcatc
     2701 cagaccaaaa gatgaattgg gtatctatta cgttagagta accccaaaca gaaacaacaa
     2761 gtcaaacttt ttccctatca gatcccataa taagatcact tacaacaact tcgaagaatc
     2821 aacttatatt gcttggtaca caaacgttga tatcgacatt ggtttgtcag aaggtcacaa
     2881 catcgaatac atccctttcg attcctatgg taacattggt tactcatggt ccaaaaaggg
     2941 taaaattttt gaaaagtaca tcaaggatgt tttgtacaaa ttgaagataa aatatgaaaa
     3001 gcaaaataac aaggtaaaaa gaaatgtcat caaaatcatc atgaactcat tgtggggtaa
     3061 attcgcccaa aagtgggtta acttcgaata cttcatcaag tcagaagatg acattgattt
     3121 cgaatccgaa gaagcatata aaatttggga tacagacttc atgttgatca aaaagattaa
     3181 ggaatctacc tacagttcta agccaataca aaacggtgtt tttactttaa gttgggccag
     3241 atatcacatg aagtctattt gggatgcagg tgccaaagaa ggtgctgaat gtatctattc
     3301 agatacagac tccatattcg ttcataagga acacttcaaa aagaatgcaa agtttatgtt
     3361 gaacggtttg aaagtaccaa tcatcggttc tgaagtcggt caattggaat tagaatgtga
     3421 atttgataaa ttgttgtgcg ccggtaaaaa gcaatacatg ggtttctaca cttacttcca
     3481 agatggtaaa ccttgcatca aggaaaagaa aagattcaaa ggtataccaa gtaactacat
     3541 catacctgaa ttgtatgctc acttgttatc tggtgcagat aaggaagcca aaattcaatt
     3601 tttaaaattc agaagagaat ggggttccgt caagggttac atagaaaata agactgtcaa
     3661 agccacctaa ctcgaggcga atttcttatg atttatgatt tttattatta aataagttat
     3721 aaaaaaaata agtgtataca aattttaaag tgactcttag gttttaaaac gaaaattctt
     3781 attcttgagt aactctttcc tgtaggtcag gttgctttct caggtatagc atgaggtcgc
     3841 tcactagtcc atggacgcgt ggtaccggcc gcgttgctgg cgtttttcca taggctcggc
     3901 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     3961 ctataaagat accaggcgtt cccccctgga agctccctcg tgcgctctcc tgttccgacc
     4021 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa
     4081 tgctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4141 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4201 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4261 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4321 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4381 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4441 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4501 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgac tgatccttca
     4561 actcagcaaa agttcgattt attcaacaaa gccacgttgt gtctcaaaat ctctgatgtt
     4621 acattgcaca agataaaaat atatcatcat gaacaataaa actgtctgct tacataaaca
     4681 gtaatacaag gggtgttatg agccatattc aacgggaaac gtcttgctcc aggccgcgat
     4741 taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat aatgtcgggc
     4801 aatcaggtgc gacaatctat cgattgtatg ggaagcccga tgcgccagag ttgtttctga
     4861 aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga ctaaactggc
     4921 tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct gatgatgcat
     4981 ggttactcac cactgcgatc cccgggaaaa cagcattcca ggtattagaa gaatatcctg
     5041 attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg cattcgattc
     5101 ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg tctcgctcag gcgcaatcac
     5161 gaatgaataa cggtttggtt gatgcgagtg attttgatga cgagcgtaat ggctggcctg
     5221 ttgaacaagt ctggaaagaa atgcataagc ttttgccatt ctcaccggat tcagtcgtca
     5281 ctcatggtga tttctcactt gataacctta tttttgacga ggggaaatta ataggttgta
     5341 ttgatgttgg acgagtcgga atcgcagacc gataccagga tcttgccatc ctatggaact
     5401 gcctcggtga gttttctcct tcattacaga aacggctttt tcaaaaatat ggtattgata
     5461 atcctgatat gaataaattg cagtttcatt tgatgctcga tgagtttttc taatcagaat
     5521 tggttaattg gttgtaacac ggtccttttc atcacgtgct ataaaaataa ttataattta
     5581 aattttttaa tataaatata taaattaaaa atagaaagta aaaaaagaaa ttaaagaaaa
     5641 aatagttttt gttttccgaa gatgtaaaag actctagggg gatcgccaac aaatactacc
     5701 ttttatcttg ctcttcctgc tctcaggtat taatgccgaa ttgtttcatc ttgtctgtgt
     5761 agaagaccac acacgaaaat cctgtgattt tacattttac ttatcgttaa tcgaatgtat
     5821 atctatttaa tctgcttttc ttgtctaata aatatatatg taaagtacgc tttttgttga
     5881 aattttttaa acctttgttt attttttttt cttcattccg taactcttct accttcttta
     5941 tttactttct aaaatccaaa tacaaaacat aaaaataaat aaacacagag taaattccca
     6001 aattattcca tcattaaaag atacgaggcg cgtgtaagtt acaggcaagc gatccgtcct
     6061 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     6121 gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     6181 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     6241 gtgttggcgg gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag
     6301 tgcaccacgc ttttcaattc aattcatcat ttttttttta ttcttttttt tgatttttta
     6361 agagcttggt gagcgctagg agtcactgcc aggtatcgtt tgaacacggc attagtcagg
     6421 gaagtcataa cacagtcctt tcccgcaatt ttctttttct attactcttg gcctcctcta
     6481 gtacactcta tattttttta tgcctcggta atgattttca tttttttttt tccacctagc
     6541 ggatgactct ttttttttct tagcgattgg cattatcaca taatgaatta tacattatat
     6601 aaagtaatgt gatttcttcg aagaatatac taaaaaatga gcaggcaaga taaacgaagg
     6661 caaagatgac agagcagaaa gccctagtaa agcgtattac aaatgaaacc aagattcaga
     6721 ttgcgatctc tttaaagggt ggtcccctag cgatagagca ctcgatcttc ccagaaaaag
     6781 aggcagaagc agtagcagaa caggccacac aatcgcaagt gattaacgtc cacacaggta
     6841 tagggtttct ggaccatatg atacatgctc tggccaagca ttccggctgg tcgctaatcg
     6901 ttgagtgcat tggtgactta cacatagacg accatcacac cactgaagac tgcgggattg
     6961 ctctcggtca agcttttaaa gaggccctag gggccgtgcg tggagtaaaa aggtttggat
     7021 caggatttgc gcctttggat gaggcacttt ccagagcggt ggttgatctt tcgaacaggc
     7081 cgtacgcagt tgtcgaactt ggtttgcaaa gggagaaagt aggtgatctc tcttgcgaga
     7141 tgatcccgca ttttcttgaa agctttgcag aggctagcag aattaccctc cacgttgatt
     7201 gtctgcgagg caagaatgat catcaccgta gtgagagtgc gttcaaggct cttgcggttg
     7261 ccataagaga agccacctcg cccaatggta ccaacgatgt tccctccacc aaaggtgttc
     7321 ttatgtagtg acaccgatta tttaaagctg ctgcatacga tatatataca tgtgtatata
     7381 tgtataccta tgaatgtcag taagtatgta tacgaacagt atgatactga agatgacaag
     7441 gtaatgcatc attctatacg tgtcattctg aacgaggcgc gctttccttt tttctttttg
     7501 ctttttcttt ttttttctct tgaactcgag aaaaaaaata tgcggtgtga aataccgcac
     7561 agatgcgtaa ggagaaaata ccgcatcagg gagctc
//
