LOCUS       sequence-445554-        7225 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Co-expression of two genes in yeast with FLAG and MYC tags and TYR1 
            selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7225)
  AUTHORS   Dong G, Nakai T, Matsuzaki T
  TITLE     A novel plasmid-based experimental system in Saccharomyces 
            cerevisiae that enables the introduction of 10 different plasmids 
            into cells.
  JOURNAL   FEBS Open Bio. 2024 Oct 10. doi: 10.1002/2211-5463.13893.
  PUBMED    39387713
REFERENCE   2  (bases 1 to 7225)
  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: "FEBS Open 
            Bio. 2024 Oct 10. doi: 10.1002/2211-5463.13893."
COMMENT     Sequence Label: pESC-TYR
FEATURES             Location/Qualifiers
     source          1..7225
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(1945..2400)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2082..2103)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2294..2313
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(2479..2644)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     2565..2589
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2792..2815)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        2862..3526
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     protein_bind    3078..3195
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3490..3513
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     3537..3556
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3537..3555
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3573..3602
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      3632..3821
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3893..3910)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4064..4652)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4144..4163)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4823..5614,5615..5683))
                     /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     5446..5465
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5684..5788)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5815..7157
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(6778..6825)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     7198..7216
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataccag cttttcaatt caattcaatt cagttatata aatatactcc ttttcacaaa
      241 agaaggccta atattatagg aaatcagcat taaaaacatt atgtatttct tttttcagcg
      301 gccgaacggt cactagaatg actcagaatg gtttttatca tctcgttgcc aatgagatta
      361 gcctctggaa gcattggctc aaagaacttt tgaacactct gaaattgctt tttgtatata
      421 tcaaaattgg caaaagaaac aaccgaactc cattctctag ccgaaataac aaattctaaa
      481 tcatctttta tgaatgattt atcatggata gctgcatcaa ttgtctgttc taataagcca
      541 ggttttaaaa aaagatattc ggacacaccc aggaatattc tgaataatgg cgtcgaacaa
      601 atcatatgat catatggatc aataccaagt tgaaaccacg aatcaacaat cgccaacaat
      661 gataaatgtg agttcggcac tggcttgcaa ttgttgctgt taccaatgct gctttttgat
      721 agcgaatatt tctctaaaat cgtgtcatcc aatagtaaga gaccagtatg ctttccaaat
      781 acaaattgct tagcttttaa tagtctatca gtaagttctt gttctttgtt atctatcatt
      841 aaactaaata gttctgttgc actggttgca tattgaagaa tttgctgatg tgcacttggg
      901 tttgttatgg ctaatcctgc gtaaacatgc cacttgttcg aatagattct tagtgatata
      961 ttaactttca cattttctag gccaccgtac catttgttta cacccagagt ccaaggataa
     1021 atctttatct ttgcccacgc agatcccata cttaagaaag cagcatgtgt cacagcttgt
     1081 gtatcagcgg taatcttgtc atgctcttca tatgtcaaat aaacttgctt acttttcaaa
     1141 catgccataa cagaattaac gaactcaaaa gattctgggt actgtgatct gtgattgata
     1201 ataacaagtg gttggccttc agtattaact ttaggcccat gaagggaatg cacggtaatg
     1261 atgtcgcagt ccttgggtaa atacttttcg aaagcctcga tttccggcag cttacaactc
     1321 gtttgacccc caacaattgt tccaacctta gaagatggtc catacgttgc gacgatctta
     1381 ctaatattgg atgcttcaac actatagata atatagtcgc tttgcctgga taccaaatga
     1441 ccatttttca ccagttcgaa tttagctgag gcatattttt ctttcagttc atcataatat
     1501 tcttccctat cacaacatat aacaccccat ccagcatctg taaatttatt agcgtataat
     1561 aggcccatat cacccagacc aattatacca atgacttttg tggctttcca ttgctcaatc
     1621 ttatcctctg ataccatgtt tatcaagtgg atatgctgtc ctctttcctt caagtgctac
     1681 ggtattcttg taattctaaa aacttttctt ggtttgaata attccagtat cactttcttc
     1741 aaagcctatt ttcatataag gcctcaaaaa aaaaaaaaaa aaaatgacac atagctgaag
     1801 tgataaactt actacactgg tataacgctt attagaggaa atctaataat aaactgttag
     1861 ctataaatag tatacaataa caatgagcgc tatatgcggt gtgaaatacc gcacagatgc
     1921 gtaaggagaa aataccgcat caggaaattg taagcgttaa tattttgtta aaattcgcgt
     1981 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     2041 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     2101 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     2161 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     2221 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     2281 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     2341 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     2401 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     2461 attacgccag ctgaattgga gcgacctcat gctatacctg agaaagcaac ctgacctaca
     2521 ggaaagagtt actcaagaat aagaattttc gttttaaaac ctaagagtca ctttaaaatt
     2581 tgtatacact tatttttttt ataacttatt taataataaa aatcataaat cataagaaat
     2641 tcgcttattt agaagtgtca acaacgtatc taccaacgat ttgacccttt tccatctttt
     2701 cgtaaatttc tggcaaggta gacaagccga caaccttgat tggagacttg accaaacctc
     2761 tggcgaagaa ttgttaatta agagctcaga tcttatcgtc gtcatccttg taatccatcg
     2821 atactagtgc ggccgccctt tagtgagggt tgaattcgaa ttttcaaaaa ttcttacttt
     2881 ttttttggat ggacgcaaag aagtttaata atcatattac atggcattac caccatatac
     2941 atatccatat acatatccat atctaatctt acttatatgt tgtggaaatg taaagagccc
     3001 cattatctta gcctaaaaaa accttctctt tggaactttc agtaatacgc ttaactgctc
     3061 attgctatat tgaagtacgg attagaagcc gccgagcggg tgacagccct ccgaaggaag
     3121 actctcctcc gtgcgtcctc gtcttcaccg gtcgcgttcc tgaaacgcag atgtgcctcg
     3181 cgccgcactg ctccgaacaa taaagattct acaatactag cttttatggt tatgaagagg
     3241 aaaaattggc agtaacctgg ccccacaaac cttcaaatga acgaatcaaa ttaacaacca
     3301 taggatgata atgcgattag ttttttagcc ttatttctgg ggtaattaat cagcgaagcg
     3361 atgatttttg atctattaac agatatataa atgcaaaaac tgcataacca ctttaactaa
     3421 tactttcaac attttcggtt tgtattactt cttattcaaa tgtaataaaa gtatcaacaa
     3481 aaaattgtta atatacctct atactttaac gtcaaggaga aaaaaccccg gatccgtaat
     3541 acgactcact atagggcccg ggcgtcgaca tggaacagaa gttgatttcc gaagaagacc
     3601 tcgagtaagc ttggtaccgc ggctagctaa gatccgctct aaccgaaaag gaaggagtta
     3661 gacaacctga agtctaggtc cctatttatt tttttatagt tatgttagta ttaagaacgt
     3721 tatttatatt tcaaattttt cttttttttc tgtacagacg cgtgtacgca tgtaacatta
     3781 tactgaaaac cttgcttgag aaggttttgg gacgctcgaa gatccagctg cattaatgaa
     3841 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     3901 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     3961 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4021 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4081 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4141 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     4201 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     4261 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     4321 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     4381 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     4441 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     4501 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     4561 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     4621 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     4681 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     4741 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     4801 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     4861 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     4921 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     4981 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5041 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     5101 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     5161 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     5221 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     5281 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     5341 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     5401 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     5461 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     5521 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     5581 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     5641 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     5701 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     5761 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgaacgaa
     5821 gcatctgtgc ttcattttgt agaacaaaaa tgcaacgcga gagcgctaat ttttcaaaca
     5881 aagaatctga gctgcatttt tacagaacag aaatgcaacg cgaaagcgct attttaccaa
     5941 cgaagaatct gtgcttcatt tttgtaaaac aaaaatgcaa cgcgagagcg ctaatttttc
     6001 aaacaaagaa tctgagctgc atttttacag aacagaaatg caacgcgaga gcgctatttt
     6061 accaacaaag aatctatact tcttttttgt tctacaaaaa tgcatcccga gagcgctatt
     6121 tttctaacaa agcatcttag attacttttt ttctcctttg tgcgctctat aatgcagtct
     6181 cttgataact ttttgcactg taggtccgtt aaggttagaa gaaggctact ttggtgtcta
     6241 ttttctcttc cataaaaaaa gcctgactcc acttcccgcg tttactgatt actagcgaag
     6301 ctgcgggtgc attttttcaa gataaaggca tccccgatta tattctatac cgatgtggat
     6361 tgcgcatact ttgtgaacag aaagtgatag cgttgatgat tcttcattgg tcagaaaatt
     6421 atgaacggtt tcttctattt tgtctctata tactacgtat aggaaatgtt tacattttcg
     6481 tattgttttc gattcactct atgaatagtt cttactacaa tttttttgtc taaagagtaa
     6541 tactagagat aaacataaaa aatgtagagg tcgagtttag atgcaagttc aaggagcgaa
     6601 aggtggatgg gtaggttata tagggatata gcacagagat atatagcaaa gagatacttt
     6661 tgagcaatgt ttgtggaagc ggtattcgca atattttagt agctcgttac agtccggtgc
     6721 gtttttggtt ttttgaaagt gcgtcttcag agcgcttttg gttttcaaaa gcgctctgaa
     6781 gttcctatac tttctagaga ataggaactt cggaatagga acttcaaagc gtttccgaaa
     6841 acgagcgctt ccgaaaatgc aacgcgagct gcgcacatac agctcactgt tcacgtcgca
     6901 cctatatctg cgtgttgcct gtatatatat atacatgaga agaacggcat agtgcgtgtt
     6961 tatgcttaaa tgcgtactta tatgcgtcta tttatgtagg atgaaaggta gtctagtacc
     7021 tcctgtgata ttatcccatt ccatgcgggg tatcgtatgc ttccttcagc actacccttt
     7081 agctgttcta tatgctgcca ctcctcaatt ggattagtct catccttcaa tgctatcatt
     7141 tcctttgata ttggatcata ctaagaaacc attattatca tgacattaac ctataaaaat
     7201 aggcgtatca cgaggccctt tcgtc
//
