LOCUS       sequence-317794-        8850 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pYES2 derivative reporter plasmid encoding PPGI1-Saccharomyces 
            cerevisiae FEX1-MFaT; as well as PGAL1-Homo sapiens 
            ADORA2A-EGFP-10HIS-MFaT..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8850)
  AUTHORS   Yoo JI, Seppala S, O'Malley MA
  TITLE     Engineered fluoride sensitivity enables biocontainment and selection
            of genetically-modified yeasts.
  JOURNAL   Nat Commun. 2020 Oct 29;11(1):5459. doi: 10.1038/s41467-020-19271-1.
  PUBMED    33122649
REFERENCE   2  (bases 1 to 8850)
  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: "2020-10-29"; volume: "11(1):5459. doi"; pages: " 
            10.1038/s41467-020-19271-1"
COMMENT     Sequence Label: pEFEX2-A2aGH
FEATURES             Location/Qualifiers
     source          1..8850
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(87..878,879..947))
                     /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     710..729
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     RBS             2067..2075
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     rep_origin      3547..4424
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     rep_origin      complement(4435..4948)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4630..4651)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4842..4861
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4970..5411
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    4970..5087
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     5382..5405
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     5443..5462
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5443..5461
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(6854..6856,6857..6859,6860..7570)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(6854..6856,6857..6859,6860..7570)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6899..6920)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(7160..7179)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7507..7528
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             7583..7612
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     terminator      7683..7930
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(7699..7717)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(8007..8024)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8178..8766)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8258..8277)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
       61 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
      121 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
      181 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
      241 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
      301 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
      361 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
      421 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
      481 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
      541 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
      601 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
      661 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
      721 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
      781 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
      841 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
      901 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact ctactcctcg
      961 tcaattctct taggattcga ttcacattca tcttttttta gctactaata ggataaatta
     1021 taggaattta taacaaattg aaatatggca ggcagcaaaa ttaaaattgt cttagttttt
     1081 tagtgtatag aagtgaatag ctatataaag tatgtgtaaa gttggtaacg gaacgaaaaa
     1141 tagaaaagga tattacatgg gaaaacatgt tgtttacgga gaaatgaaaa gtatattgta
     1201 ttttgtacga gctaaaagta cagtgggaac aaagtcgatt ttgttacatc tacactgttg
     1261 ttatcagatc tcgagctatt acaagtcttc ttcagaaata agcttttgtt atcaggagac
     1321 ggtgactgag gttccttgac cccagtagtc cataccatag taagaacccg tacagtaata
     1381 gatacccatg tcttcaactc ttaagctaac aaatcgggtt ggtcaatcct ctagtccaag
     1441 cataggatcc gagagttatc accaataagc aatatgaaat tgcaatcgaa acagtataat
     1501 aaataagcat gttaataaat gatagcttat acccttcatt gataaacgtg ctaatggtac
     1561 tcaaagtgcc gcaaaatccg gatattaacg cagaaacaat atgacatgaa ttcaacgagt
     1621 tcacaattgg aacgtccgta gaaaagtgtt ttttaccacg ttgcaccatg gtaaatatgc
     1681 caatcaataa agtggcgaaa acgtttgcca agaaagtacc caatggaaac tttttgtttg
     1741 ttttattaaa catttctgcc aaccaatatc ttaggaaacc agcgaaaatt ccaaatagac
     1801 atggtagtgt ccacttgccc ctcgaatagt tctcataata cgcacacagc accacaaaca
     1861 aaataattaa tggaaaagct agtgcatacg cgagtttatc cacaatatca aaaaatttct
     1921 taaaaaatgg tgctggagtt ttgaatttaa actctagaat atttttttcc atttcttgtg
     1981 tatcaacatc accagcattt tcctttacag tgtctgaggg aggggtatat ggcttcgaaa
     2041 acgaactaga tccataagca acaataacct ccttaccaag ttgcctaccg aaaataagac
     2101 tacccatgga aaccataaga tgaacgagca agacagataa aaactccatt ataccgtaag
     2161 ctctgttcgg cagttttgtg tggttagcga tattaccgtt agttaagttt gtagagtgtt
     2221 cgaacatctc tagcagcata ctagaaaagg aagacagggc accacagtac ccagtagtta
     2281 cacctaaaaa caagacttgg tgatctttca tccaagtgta agcatttaaa gattgcataa
     2341 tccccattag catgcaggat gaacaattag accagagaac ggtcggagca gtaacgaagg
     2401 ccggagtgta ggtggaaagc gccgttatag cctgacgtgt ttcagtacct aatatgcaga
     2461 acgtggtgaa tgtacaaaaa acatgaatat aatggctcaa cttgtgattt gatatgaccg
     2521 gattgaaaat cattttgaat tctttttagg ctggtatctt gattctaaat cgattttgca
     2581 agactagagg aatatgtatt aaaattcact gaatgcaagt aaaatattaa gtaaaagaaa
     2641 gcattgatta tgaagaaagg acctttttaa gatatgagaa caatgagaaa aggaaaaatg
     2701 ggacgaaaca aatagggaaa attccgtctg taaaaccaat tgtagtaaca aaaaacgctc
     2761 atttcaatag caaaaaaata gaataccgtc aatggatata tatagacacg ggacattata
     2821 acaacgtgca gatggtgatg attgcaaacc caaaaaatta ccagcttcct gtaattccag
     2881 gttgagcgcg tggcttccta taatccaata cacccacacc cacccagatc gtgatttttg
     2941 ttacaggtca gctcttatcg atgataagct gtcaaacatg agaattaatt ccacggacta
     3001 tagactatac ctagtatact ccgtctactg tacgatacac ttccgctcag gtccttgtcc
     3061 tttaacgagg ccttaccact cttttgttac tctattgatc cagctcagca aaggcagtgt
     3121 gatctaagat tctatcttcg cgatgtagta aaactagcta gaccgagaaa gagactagaa
     3181 atgcaaaagg cacttctaca atggctgcca tcattattat ccgatgtgac gctgcagctt
     3241 ctcaatgata ttcgaatacg ctttgaggag atacagccta atatccgaca aactgtttta
     3301 cagatttacg atcgtacttg ttacccatca ttgaattttg aacatccgaa cctgggagtt
     3361 ttccctgaaa cagatagtat atttgaacct gtataataat atatagtcta gcgctttacg
     3421 gaagacaatg tatgtatttc ggttcctgga gaaactattg catctattgc ataggtaatc
     3481 ttgcacgtcg catccccggt tcattttctg cgtttccatc ttgcacttca atagcatatc
     3541 tttgttaacg aagcatctgt gcttcatttt gtagaacaaa aatgcaacgc gagagcgcta
     3601 atttttcaaa caaagaatct gagctgcatt tttacagaac agaaatgcaa cgcgaaagcg
     3661 ctattttacc aacgaagaat ctgtgcttca tttttgtaaa acaaaaatgc aacgcgagag
     3721 cgctaatttt tcaaacaaag aatctgagct gcatttttac agaacagaaa tgcaacgcga
     3781 gagcgctatt ttaccaacaa agaatctata cttctttttt gttctacaaa aatgcatccc
     3841 gagagcgcta tttttctaac aaagcatctt agattacttt ttttctcctt tgtgcgctct
     3901 ataatgcagt ctcttgataa ctttttgcac tgtaggtccg ttaaggttag aagaaggcta
     3961 ctttggtgtc tattttctct tccataaaaa aagcctgact ccacttcccg cgtttactga
     4021 ttactagcga agctgcgggt gcattttttc aagataaagg catccccgat tatattctat
     4081 accgatgtgg attgcgcata ctttgtgaac agaaagtgat agcgttgatg attcttcatt
     4141 ggtcagaaaa ttatgaacgg tttcttctat tttgtctcta tatactacgt ataggaaatg
     4201 tttacatttt cgtattgttt tcgattcact ctatgaatag ttcttactac aatttttttg
     4261 tctaaagagt aatactagag ataaacataa aaaatgtaga ggtcgagttt agatgcaagt
     4321 tcaaggagcg aaaggtggat gggtaggtta tatagggata tagcacagag atatatagca
     4381 aagagatact tttgagcaat gtttgtggaa gcggtattcg caatgggaag ctccaccccg
     4441 gttgataatc agaaaagccc caaaaacagg aagattgtat aagcaaatat ttaaattgta
     4501 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     4561 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     4621 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     4681 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     4741 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     4801 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     4861 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     4921 gcgcttaatg cgccgctaca gggcgcgtgg ggatcgatcc actagtccac ggattagaag
     4981 ccgccgagcg ggtgacagcc ctccgaagga agactctcct ccgtgcgtcc tcgtcttcac
     5041 cggtcgcgtt cctgaaacgc agatgtgcct cgcgccgcac tgctccgaac aataaagatt
     5101 ctacaatact agcttttatg gttatgaaga ggaaaaattg gcagtaacct ggccccacaa
     5161 accttcaaat gaacgaatca aattaacaac cataggatga taatgcgatt agttttttag
     5221 ccttatttct ggggtaatta atcagcgaag cgatgatttt tgatctatta acagatatat
     5281 aaatgcaaaa actgcataac cactttaact aatactttca acattttcgg tttgtattac
     5341 ttcttattca aatgtaataa aagtatcaac aaaaaattgt taatatacct ctatacttta
     5401 acgtcaagga gaaaaaaccc cggatcggac tactagcagc tgtaatacga ctcactatag
     5461 ggaatattaa gcttggatca tgaaggtttt gattgtcttg ttggctatct tcgctgcttt
     5521 gcccttggct ctagctcaac cggttatttc tactaccgtc ggttccgctg cagaaggctc
     5581 tttggacaag agagaagctc ggccggacgt catgcccatc atgggctcct cggtgtacat
     5641 cacggtggag ctggccattg ctgtgctggc catcctgggc aatgtgctgg tgtgctgggc
     5701 cgtgtggctc aacagcaacc tgcagaacgt caccaactac tttgtggtgt cactggcggc
     5761 ggccgacatc gcagtgggtg tgctcgccat cccctttgcc atcaccatca gcaccgggtt
     5821 ctgcgctgcc tgccacggct gcctcttcat tgcctgcttc gtcctggtcc tcacgcagag
     5881 ctccatcttc agtctcctgg ccatcgccat tgaccgctac attgccatcc gcatcccgct
     5941 ccggtacaat ggcttggtga ccggcacgag ggctaagggc atcattgcca tctgctgggt
     6001 gctgtcgttt gccatcggcc tgactcccat gctaggttgg aacaactgcg gtcagccaaa
     6061 ggagggcaag aaccactccc agggctgcgg ggagggccaa gtggcctgtc tctttgagga
     6121 tgtggtcccc atgaactaca tggtgtactt caacttcttt gcctgtgtgc tggtgcccct
     6181 gctgctcatg ctgggtgtct atttgcggat cttcctggcg gcgcgacgac agctgaagca
     6241 gatggagagc cagcctctgc cgggggagcg ggcacggtcc acactgcaga aggaggtcca
     6301 tgctgccaag tcactggcca tcattgtggg gctctttgcc ctctgctggc tgcccctaca
     6361 catcatcaac tgcttcactt tcttctgccc cgactgcagc cacgcccctc tctggctcat
     6421 gtacctggcc atcgtcctct cccacaccaa ttcggttgtg aatcccttca tctacgccta
     6481 ccgtatccgc gagttccgcc agaccttccg caagatcatt cgcagccacg tcctgaggca
     6541 gcaagaacct ttcaaggcag ctggcaccag tgcccgggtc ttggcagctc atggcagtga
     6601 cggagagcag gtcagcctcc gtctcaacgg ccacccgcca ggagtgtggg ccaacggcag
     6661 tgctccccac cctgagcgga ggcccaatgg ctacgccctg gggctggtga gtggagggag
     6721 tgcccaagag tcccagggga acacgggcct cccagacgtg gagctcctta gccatgagct
     6781 caagggagtg tgcccagagc cccctggcct agatgacccc ctggcccagg atggagcagg
     6841 agtgtccccg cggatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     6901 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     6961 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     7021 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     7081 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     7141 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     7201 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     7261 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     7321 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     7381 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     7441 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     7501 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     7561 gctgtacaag cacgtgtcat cacatcatca tcatcatcat catcatcatc attaataact
     7621 taagagttga agacatgggt atctattact gtacggctcg agcatgcatc tagagggccg
     7681 catcatgtaa ttagttatgt cacgcttaca ttcacgccct ccccccacat ccgctctaac
     7741 cgaaaaggaa ggagttagac aacctgaagt ctaggtccct atttattttt ttatagttat
     7801 gttagtatta agaacgttat ttatatttca aatttttctt ttttttctgt acagacgcgt
     7861 gtacgcatgt aacattatac tgaaaacctt gcttgagaag gttttgggac gctcgaaggc
     7921 tttaatttgc aagctgcggc cctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     7981 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     8041 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     8101 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     8161 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     8221 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     8281 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     8341 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     8401 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     8461 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     8521 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcag tgctacagag
     8581 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     8641 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     8701 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     8761 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     8821 cgttaaggga ttttggtcat gagattatca
//
