LOCUS       sequence-270188-        9051 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCS4333 CEN/ARS vector, PGAL1-yeTadA1.0-100AA linker-T7 RNAP-TCYC1, 
            TRP1 selectable marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9051)
  AUTHORS   Cravens A, Jamil OK, Kong D, Sockolosky JT, Smolke CD
  TITLE     Polymerase-guided base editing enables in vivo mutagenesis and rapid
            protein engineering.
  JOURNAL   Nat Commun. 2021 Mar 11;12(1):1579. doi: 10.1038/s41467-021-21876-z.
  PUBMED    33707425
REFERENCE   2  (bases 1 to 9051)
  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: "Nat 
            Commun."; date: "2021-03-11"; volume: "12(1):1579. doi"; pages: " 
            10.1038/s41467-021-21876-z"
COMMENT     Sequence Label: pCS4333 yeTadA1.0-T7 RNAP
FEATURES             Location/Qualifiers
     source          1..9051
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     69..88
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        109..389
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             390..1064
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     rep_origin      complement(1167..1622)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1304..1325)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1516..1535
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(1701..1866)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     1787..1811
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2014..2037)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        2084..2748
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     protein_bind    2300..2417
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     2712..2735
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     terminator      6205..6394
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(6466..6483)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6637..7225)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6717..6736)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7396..8187,8188..8256))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSHIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8019..8038
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8257..8361)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    8398..8901
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     8942..8960
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(8998..9020)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
       61 taactatgcg gcatcagagc agattgtact gagagtgcac catagatcac gacattacta
      121 tatatataat ataggaagca tttaatagaa cagcatcgta atatatgtgt actttgcagt
      181 tatgacgcca gatggcagta gtggaagata ttctttattg aaaaatagct tgtcacctta
      241 cgtacaatct tgatccggag cttttctttt tttgccgatt aagaattaat tcggtcgaaa
      301 aaagaaaagg agagggccaa gagggagggc attggtgact attgagcacg tgagtatacg
      361 tgattaagca cacaaaggca gcttggagta tgtctgttat taatttcaca ggtagttctg
      421 gtccattggt gaaagtttgc ggcttgcaga gcacagaggc cgcagaatgt gctctagatt
      481 ccgatgctga cttgctgggt attatatgtg tgcccaatag aaagagaaca attgacccgg
      541 ttattgcaag gaaaatttca agtcttgtaa aagcatataa aaatagttca ggcactccga
      601 aatacttggt tggcgtgttt cgtaatcaac ctaaggagga tgttttggct ctggtcaatg
      661 attacggcat tgatatcgtc caactgcatg gagatgagtc gtggcaagaa taccaagagt
      721 tcctcggttt gccagttatt aaaagactcg tatttccaaa agactgcaac atactactca
      781 gtgcagcttc acagaaacct cattcgttta ttcccttgtt tgattcagaa gcaggtggga
      841 caggtgaact tttggattgg aactcgattt ctgactgggt tggaaggcaa gagagccccg
      901 aaagcttaca ttttatgtta gctggtggac tgacgccaga aaatgttggt gatgcgctta
      961 gattaaatgg cgttattggt gttgatgtaa gcggaggtgt ggagacaaat ggtgtaaaag
     1021 actctaacaa aatagcaaat ttcgtcaaaa atgctaagaa ataggttatt actgagtagt
     1081 atttatttaa gtattgtttg tgcacttgcc gatctatgcg gtgtgaaata ccgcacagat
     1141 gcgtaaggag aaaataccgc atcaggaaat tgtaagcgtt aatattttgt taaaattcgc
     1201 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     1261 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     1321 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     1381 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     1441 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     1501 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     1561 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     1621 gtccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     1681 ctattacgcc agctgaattg gagcgacctc atgctatacc tgagaaagca acctgaccta
     1741 caggaaagag ttactcaaga ataagaattt tcgttttaaa acctaagagt cactttaaaa
     1801 tttgtataca cttatttttt ttataactta tttaataata aaaatcataa atcataagaa
     1861 attcgcttat ttagaagtgt caacaacgta tctaccaacg atttgaccct tttccatctt
     1921 ttcgtaaatt tctggcaagg tagacaagcc gacaaccttg attggagact tgaccaaacc
     1981 tctggcgaag aattgttaat taagagctca gatcttatcg tcgtcatcct tgtaatccat
     2041 cgatactagt gcggccgccc tttagtgagg gttgaattcg aattttcaaa aattcttact
     2101 ttttttttgg atggacgcaa agaagtttaa taatcatatt acatggcatt accaccatat
     2161 acatatccat atacatatcc atatctaatc ttacttatat gttgtggaaa tgtaaagagc
     2221 cccattatct tagcctaaaa aaaccttctc tttggaactt tcagtaatac gcttaactgc
     2281 tcattgctat attgaagtac ggattagaag ccgccgagcg ggtgacagcc ctccgaagga
     2341 agactctcct ccgtgcgtcc tcgtcttcac cggtcgcgtt cctgaaacgc agatgtgcct
     2401 cgcgccgcac tgctccgaac aataaagatt ctacaatact agcttttatg gttatgaaga
     2461 ggaaaaattg gcagtaacct ggccccacaa accttcaaat gaacgaatca aattaacaac
     2521 cataggatga taatgcgatt agttttttag ccttatttct ggggtaatta atcagcgaag
     2581 cgatgatttt tgatctatta acagatatat aaatgcaaaa actgcataac cactttaact
     2641 aatactttca acattttcgg tttgtattac ttcttattca aatgtaataa aagtatcaac
     2701 aaaaaattgt taatatacct ctatacttta acgtcaagga gaaaaaaccc cggatccgat
     2761 gtccgaggtt gagttcagcc atgagtactg gatgagacac gcattgactc tagcaaagag
     2821 ggcgagagac gagagggaag tccccgtcgg cgccgtatta gtcttaaata atagggtaat
     2881 tggtgaaggg tggaaccgtg ctattgggtt acatgacccg acagctcacg cggagattat
     2941 ggcacttaga caaggcggtc ttgtaatgca gaactaccgt ttaatagacg caacactgta
     3001 cgtcacattc gaaccttgtg taatgtgtgc cggagccatg atccactccc gtataggccg
     3061 tgttgtcttt ggggtcagaa acgcgaagac aggagcagcc ggttccctga tggatgtgtt
     3121 acactaccct ggaatgaatc atagtgtcga gataacggag ggtatcctgg cagacgagtg
     3181 tgccgcgttg ctgtgttatt tttttaggat gccccgtcgg gtctttaatg cgcagaaaaa
     3241 agcgcagccc tctaccggct ctggaggtgg aggttctgca gagtatgtgc gggccctctt
     3301 tgactttaat gggaatgatg aagaagatct tccctttaag aaaggagaca tcctgagaat
     3361 ccgggataag cctgaagagc agtggtggaa tgcagaggac agcgaaggaa agagggggat
     3421 gattcctgtc ccttacgtgg agaagtattc cggagactat aaggaccacg acggagacta
     3481 caaggatcat gatattgatt acaaagacga tgacaatcag gttaacacga ttaacatagc
     3541 taagaacgac ttctctgaca tcgaactggc tgctatcccg ttcaacactc tggctgacca
     3601 ttacggtgag cgtttagctc gcgaacagtt ggcccttgag catgagtctt acgagatggg
     3661 tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa gctggtgagg ttgcggataa
     3721 cgctgccgcc aagcctctca tcactaccct actccctaag atgattgcac gcatcaacga
     3781 ctggtttgag gaagtgaaag ctaagcgcgg caagcgcccg acagccttcc agttcctgca
     3841 agaaatcaag ccgggagccg tagcgtacat caccattaag accactctgg cttgcctaac
     3901 cagtgctgac aatacaaccg ttcaggctgt agcaagcgca atcggtcggg ccattgagga
     3961 cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag cacttcaaga aaaacgttga
     4021 ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa gcatttatgc aagttgtcga
     4081 ggctgacatg ctctctaagg gtctactcgg tggcgaggcg tggtcttcgt ggcataagga
     4141 agactctatt catgtaggag tacgctgcat cgagatgctc attgagtcaa ccggaatggt
     4201 tagcttacac cgccaaaatg ctggcgtagt aggtcaagac tctgagacta tcgaactcgc
     4261 acctgaatac gctgaggcta tagcaacccg tgcaggtgcg ctggctggca tctctccgat
     4321 gttccaacct tgcgtagttc ctcctaagcc gtggactggc attactggtg gtggctattg
     4381 ggctaacggt cgtcgtcctc tggcgctggt gcgtactcac agtaagaaag cactgatgcg
     4441 ctacgaagac gtttacatgc ctgaggtgta caaagcgatt aacattgcgc aaaacaccgc
     4501 atggaaaatc aacaagaaag tcctagcggt cgccaacgta atcaccaagt ggaagcattg
     4561 tccggtcgag gacatccctg cgattgagcg tgaagaactc ccgatgaaac cggaagacat
     4621 cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct gccgctgctg tgtaccgcaa
     4681 ggacaaggct cgcaagtctc gccgtatcag ccttgagttc atgcttgagc aagccaataa
     4741 gtttgctaac cataaggcca tctggttccc ttacaacatg gactggcgcg gtcgtgttta
     4801 cgctgtgtca atgttcaacc cgcaaggtaa cgatatgacc aaaggactgc ttacgctggc
     4861 gaaaggtaaa ccaatcggta aggaaggtta ctactggctg aaaatccacg gtgcaaactg
     4921 tgcgggtgtc gataaggttc cgttccctga gcgcatcaag ttcattgagg aaaaccacga
     4981 gaacatcatg gcttgcgcta agtctccact ggagaacact tggtgggctg agcaagattc
     5041 tccgttctgc ttccttgcgt tctgctttga gtacgctggg gtacagcacc acggcctgag
     5101 ctataactgc tcccttccgc tggcgtttga cgggtcttgc tctggcatcc agcacttctc
     5161 cgcgatgctc cgagatgagg taggtggtcg cgcggttaac ttgcttccta gtgaaaccgt
     5221 tcaggacatc tacgggattg ttgctaagaa agtcaacgag attctacaag cagacgcaat
     5281 caatgggacc gataacgaag tagttaccgt gaccgatgag aacactggtg aaatctctga
     5341 gaaagtcaag ctgggcacta aggcactggc tggtcaatgg ctggcttacg gtgttactcg
     5401 cagtgtgact aagcgttcag tcatgacgct ggcttacggg tccaaagagt tcggcttccg
     5461 tcaacaagtg ctggaagata ccattcagcc agctattgat tccggcaagg gtctgatgtt
     5521 cactcagccg aatcaggctg ctggatacat ggctaagctg atttgggaat ctgtgagcgt
     5581 gacggtggta gctgcggttg aagcaatgaa ctggcttaag tctgctgcta agctgctggc
     5641 tgctgaggtc aaagataaga agactggaga gattcttcgt aagcgttgcg ctgtgcattg
     5701 ggtaactcct gatggtttcc ctgtgtggca ggaatacaag aagcctattc agacgcgctt
     5761 gaacctgatg ttcctcggtc agttccgctt acagcctacc attaacacca acaaagatag
     5821 cgagattgat gcacacaaac aggagtctgg tatcgctcct aactttgtac acagccaaga
     5881 cggtagccac cttcgtaaga ctgtagtgtg ggcacacgag aagtacggaa tcgaatcttt
     5941 tgcactgatt cacgactcct tcggtaccat tccggctgac gctgcgaacc tgttcaaagc
     6001 agtgcgcgaa actatggttg acacatatga gtcttgtgat gtactggctg atttctacga
     6061 ccagttcgct gaccagttgc acgagtctca attggacaaa atgccagcac ttccggctaa
     6121 aggtaacttg aacctccgtg acatcttaga gtcggacttc gcgttcgcgt aaaatcagta
     6181 agcttggtac cgcggctagc taagatccgc tctaaccgaa aaggaaggag ttagacaacc
     6241 tgaagtctag gtccctattt atttttttat agttatgtta gtattaagaa cgttatttat
     6301 atttcaaatt tttctttttt ttctgtacag acgcgtgtac gcatgtaaca ttatactgaa
     6361 aaccttgctt gagaaggttt tgggacgctc gaagatccag ctgcattaat gaatcggcca
     6421 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     6481 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     6541 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     6601 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     6661 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     6721 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     6781 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     6841 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     6901 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     6961 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     7021 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     7081 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     7141 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     7201 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     7261 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     7321 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     7381 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     7441 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tatgggaggg
     7501 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     7561 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     7621 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     7681 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     7741 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     7801 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     7861 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     7921 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     7981 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     8041 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     8101 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     8161 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     8221 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     8281 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     8341 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgggt ccttttcatc
     8401 acgtgctata aaaataatta taatttaaat tttttaatat aaatatataa attaaaaata
     8461 gaaagtaaaa aaagaaatta aagaaaaaat agtttttgtt ttccgaagat gtaaaagact
     8521 ctagggggat cgccaacaaa tactaccttt tatcttgctc ttcctgctct caggtattaa
     8581 tgccgaattg tttcatcttg tctgtgtaga agaccacaca cgaaaatcct gtgattttac
     8641 attttactta tcgttaatcg aatgtatatc tatttaatct gcttttcttg tctaataaat
     8701 atatatgtaa agtacgcttt ttgttgaaat tttttaaacc tttgtttatt tttttttctt
     8761 cattccgtaa ctcttctacc ttctttattt actttctaaa atccaaatac aaaacataaa
     8821 aataaataaa cacagagtaa attcccaaat tattccatca ttaaaagata cgaggcgcgt
     8881 gtaagttaca ggcaagcgat ccgtctaaga aaccattatt atcatgacat taacctataa
     8941 aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct
     9001 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg c
//
