LOCUS       sequence-445562-        7246 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Co-expression of two genes in yeast with V5 and HA tags and TYR1 
            selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7246)
  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 7246)
  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 (V5 and HA)
FEATURES             Location/Qualifiers
     source          1..7246
                     /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(2789..2830)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     promoter        2883..3547
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     protein_bind    3099..3216
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3511..3534
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     3558..3577
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3558..3576
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3600..3626
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     terminator      3653..3842
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3914..3931)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4085..4673)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4165..4184)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4844..5635,5636..5704))
                     /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     5467..5486
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5705..5809)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5836..7178
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(6799..6846)
                     /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     7219..7237
                     /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 agagctcagg tagaatccag acccagcaga gggttaggga
     2821 taggcttacc agatctcatc gatactagtg cggccgccct ttagtgaggg ttgaattcga
     2881 attttcaaaa attcttactt tttttttgga tggacgcaaa gaagtttaat aatcatatta
     2941 catggcatta ccaccatata catatccata tacatatcca tatctaatct tacttatatg
     3001 ttgtggaaat gtaaagagcc ccattatctt agcctaaaaa aaccttctct ttggaacttt
     3061 cagtaatacg cttaactgct cattgctata ttgaagtacg gattagaagc cgccgagcgg
     3121 gtgacagccc tccgaaggaa gactctcctc cgtgcgtcct cgtcttcacc ggtcgcgttc
     3181 ctgaaacgca gatgtgcctc gcgccgcact gctccgaaca ataaagattc tacaatacta
     3241 gcttttatgg ttatgaagag gaaaaattgg cagtaacctg gccccacaaa ccttcaaatg
     3301 aacgaatcaa attaacaacc ataggatgat aatgcgatta gttttttagc cttatttctg
     3361 gggtaattaa tcagcgaagc gatgattttt gatctattaa cagatatata aatgcaaaaa
     3421 ctgcataacc actttaacta atactttcaa cattttcggt ttgtattact tcttattcaa
     3481 atgtaataaa agtatcaaca aaaaattgtt aatatacctc tatactttaa cgtcaaggag
     3541 aaaaaacccc ggatccgtaa tacgactcac tatagggccc gggcgtcgac atgctcgagt
     3601 acccatacga tgttccagat tacgcttaag cttggtaccg cggctagcta agatccgctc
     3661 taaccgaaaa ggaaggagtt agacaacctg aagtctaggt ccctatttat ttttttatag
     3721 ttatgttagt attaagaacg ttatttatat ttcaaatttt tctttttttt ctgtacagac
     3781 gcgtgtacgc atgtaacatt atactgaaaa ccttgcttga gaaggttttg ggacgctcga
     3841 agatccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     3901 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     3961 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4021 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4081 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4141 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4201 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4261 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4321 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4381 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4441 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4501 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     4561 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     4621 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     4681 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     4741 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     4801 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     4861 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     4921 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     4981 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5041 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     5101 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     5161 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     5221 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     5281 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     5341 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     5401 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     5461 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     5521 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     5581 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     5641 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     5701 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     5761 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     5821 gaaaagtgcc acctgaacga agcatctgtg cttcattttg tagaacaaaa atgcaacgcg
     5881 agagcgctaa tttttcaaac aaagaatctg agctgcattt ttacagaaca gaaatgcaac
     5941 gcgaaagcgc tattttacca acgaagaatc tgtgcttcat ttttgtaaaa caaaaatgca
     6001 acgcgagagc gctaattttt caaacaaaga atctgagctg catttttaca gaacagaaat
     6061 gcaacgcgag agcgctattt taccaacaaa gaatctatac ttcttttttg ttctacaaaa
     6121 atgcatcccg agagcgctat ttttctaaca aagcatctta gattactttt tttctccttt
     6181 gtgcgctcta taatgcagtc tcttgataac tttttgcact gtaggtccgt taaggttaga
     6241 agaaggctac tttggtgtct attttctctt ccataaaaaa agcctgactc cacttcccgc
     6301 gtttactgat tactagcgaa gctgcgggtg cattttttca agataaaggc atccccgatt
     6361 atattctata ccgatgtgga ttgcgcatac tttgtgaaca gaaagtgata gcgttgatga
     6421 ttcttcattg gtcagaaaat tatgaacggt ttcttctatt ttgtctctat atactacgta
     6481 taggaaatgt ttacattttc gtattgtttt cgattcactc tatgaatagt tcttactaca
     6541 atttttttgt ctaaagagta atactagaga taaacataaa aaatgtagag gtcgagttta
     6601 gatgcaagtt caaggagcga aaggtggatg ggtaggttat atagggatat agcacagaga
     6661 tatatagcaa agagatactt ttgagcaatg tttgtggaag cggtattcgc aatattttag
     6721 tagctcgtta cagtccggtg cgtttttggt tttttgaaag tgcgtcttca gagcgctttt
     6781 ggttttcaaa agcgctctga agttcctata ctttctagag aataggaact tcggaatagg
     6841 aacttcaaag cgtttccgaa aacgagcgct tccgaaaatg caacgcgagc tgcgcacata
     6901 cagctcactg ttcacgtcgc acctatatct gcgtgttgcc tgtatatata tatacatgag
     6961 aagaacggca tagtgcgtgt ttatgcttaa atgcgtactt atatgcgtct atttatgtag
     7021 gatgaaaggt agtctagtac ctcctgtgat attatcccat tccatgcggg gtatcgtatg
     7081 cttccttcag cactaccctt tagctgttct atatgctgcc actcctcaat tggattagtc
     7141 tcatccttca atgctatcat ttcctttgat attggatcat actaagaaac cattattatc
     7201 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtc
//
