LOCUS       sequence-317846-        9148 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pYES2 derivative reporter plasmid encoding PREV1-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 9148)
  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 9148)
  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: pEFEX3-A2aGH
FEATURES             Location/Qualifiers
     source          1..9148
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(129..920,921..989))
                     /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     752..771
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     RBS             2109..2117
                     /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      3887..4764
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     rep_origin      complement(4775..5288)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4970..4991)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5182..5201
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5310..5751
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    5310..5427
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     5722..5745
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     5783..5802
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5783..5801
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(7194..7196,7197..7199,7200..7910)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(7194..7196,7197..7199,7200..7910)
                     /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(7239..7260)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(7500..7519)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7847..7868
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             7923..7952
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     terminator      8023..8270
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(8039..8057)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(8347..8364)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8518..9106)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8598..8617)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
       61 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
      121 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
      181 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
      241 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
      301 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
      361 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
      421 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
      481 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
      541 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
      601 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
      661 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
      721 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
      781 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
      841 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
      901 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
      961 agggcgacac ggaaatgttg aatactcata ctctactcct cgtcaattct cttaggattc
     1021 gattcacatt catctttttt tagctactaa taggataaat tataggaatt tataacaaat
     1081 tgaaatatgg caggcagcaa aattaaaatt gtcttagttt tttagtgtat agaagtgaat
     1141 agctatataa agtatgtgta aagttggtaa cggaacgaaa aatagaaaag gatattacat
     1201 gggaaaacat gttgtttacg gagaaatgaa aagtatattg tattttgtac gagctaaaag
     1261 tacagtggga acaaagtcga ttttgttaca tctacactgt tgttatcaga tctcgagcta
     1321 ttacaagtct tcttcagaaa taagcttttg ttatcaggag acggtgactg aggttccttg
     1381 accccagtag tccataccat agtaagaacc cgtacagtaa tagataccca tgtcttcaac
     1441 tcttaagcta acaaatcggg ttggtcaatc ctctagtcca agcataggat ccgagagtta
     1501 tcaccaataa gcaatatgaa attgcaatcg aaacagtata ataaataagc atgttaataa
     1561 atgatagctt atacccttca ttgataaacg tgctaatggt actcaaagtg ccgcaaaatc
     1621 cggatattaa cgcagaaaca atatgacatg aattcaacga gttcacaatt ggaacgtccg
     1681 tagaaaagtg ttttttacca cgttgcacca tggtaaatat gccaatcaat aaagtggcga
     1741 aaacgtttgc caagaaagta cccaatggaa actttttgtt tgttttatta aacatttctg
     1801 ccaaccaata tcttaggaaa ccagcgaaaa ttccaaatag acatggtagt gtccacttgc
     1861 ccctcgaata gttctcataa tacgcacaca gcaccacaaa caaaataatt aatggaaaag
     1921 ctagtgcata cgcgagttta tccacaatat caaaaaattt cttaaaaaat ggtgctggag
     1981 ttttgaattt aaactctaga atattttttt ccatttcttg tgtatcaaca tcaccagcat
     2041 tttcctttac agtgtctgag ggaggggtat atggcttcga aaacgaacta gatccataag
     2101 caacaataac ctccttacca agttgcctac cgaaaataag actacccatg gaaaccataa
     2161 gatgaacgag caagacagat aaaaactcca ttataccgta agctctgttc ggcagttttg
     2221 tgtggttagc gatattaccg ttagttaagt ttgtagagtg ttcgaacatc tctagcagca
     2281 tactagaaaa ggaagacagg gcaccacagt acccagtagt tacacctaaa aacaagactt
     2341 ggtgatcttt catccaagtg taagcattta aagattgcat aatccccatt agcatgcagg
     2401 atgaacaatt agaccagaga acggtcggag cagtaacgaa ggccggagtg taggtggaaa
     2461 gcgccgttat agcctgacgt gtttcagtac ctaatatgca gaacgtggtg aatgtacaaa
     2521 aaacatgaat ataatggctc aacttgtgat ttgatatgac cggattgaaa atcattttga
     2581 attctcgctg gatatgccta gaaatgcagt atcgatttat tttgagaaaa tctgtcaact
     2641 cgttacttct ttgataggga aatctttagt taaatttgtt taaaattaag aaccaggtct
     2701 ttagactatg ccaaaaggta ttggtgctta aaggttgccg taaactctaa ctctatatat
     2761 ctctaagcca atgtttaagt tgatgcgatg tgcatgatca cttcacattt ttttcatgat
     2821 tttgttttct tccttagcta cttttttccg tcttggtctg tgtaaaaaaa aaatcgcttc
     2881 actgaagaaa aatataaata aaaaaaggac atcagcaagt tcgtagtcaa taaatgtgat
     2941 gagtaatttg cgataatcta ccccaactca aagcattcaa ttcaaatgaa agaagagttc
     3001 gtgaatttta atctggctta cagattggat catagcttct aatatatctt cctttttatt
     3061 ttaaagtgat taaaatatgt ggatgtctgc attaaggtga aggcagacta tagtgaaaaa
     3121 tagtagcaaa ggcttcatat aatgaactac ctagttgtat tcaattggaa tctgtgaacg
     3181 cggcattaaa agaatatttg agcttttttt gattgtatta acctgaagct atcacggttt
     3241 agactcatta aaagaaaaat atccggttgt atcggataac acacaggtca gctcttatcg
     3301 atgataagct gtcaaacatg agaattaatt ccacggacta tagactatac ctagtatact
     3361 ccgtctactg tacgatacac ttccgctcag gtccttgtcc tttaacgagg ccttaccact
     3421 cttttgttac tctattgatc cagctcagca aaggcagtgt gatctaagat tctatcttcg
     3481 cgatgtagta aaactagcta gaccgagaaa gagactagaa atgcaaaagg cacttctaca
     3541 atggctgcca tcattattat ccgatgtgac gctgcagctt ctcaatgata ttcgaatacg
     3601 ctttgaggag atacagccta atatccgaca aactgtttta cagatttacg atcgtacttg
     3661 ttacccatca ttgaattttg aacatccgaa cctgggagtt ttccctgaaa cagatagtat
     3721 atttgaacct gtataataat atatagtcta gcgctttacg gaagacaatg tatgtatttc
     3781 ggttcctgga gaaactattg catctattgc ataggtaatc ttgcacgtcg catccccggt
     3841 tcattttctg cgtttccatc ttgcacttca atagcatatc tttgttaacg aagcatctgt
     3901 gcttcatttt gtagaacaaa aatgcaacgc gagagcgcta atttttcaaa caaagaatct
     3961 gagctgcatt tttacagaac agaaatgcaa cgcgaaagcg ctattttacc aacgaagaat
     4021 ctgtgcttca tttttgtaaa acaaaaatgc aacgcgagag cgctaatttt tcaaacaaag
     4081 aatctgagct gcatttttac agaacagaaa tgcaacgcga gagcgctatt ttaccaacaa
     4141 agaatctata cttctttttt gttctacaaa aatgcatccc gagagcgcta tttttctaac
     4201 aaagcatctt agattacttt ttttctcctt tgtgcgctct ataatgcagt ctcttgataa
     4261 ctttttgcac tgtaggtccg ttaaggttag aagaaggcta ctttggtgtc tattttctct
     4321 tccataaaaa aagcctgact ccacttcccg cgtttactga ttactagcga agctgcgggt
     4381 gcattttttc aagataaagg catccccgat tatattctat accgatgtgg attgcgcata
     4441 ctttgtgaac agaaagtgat agcgttgatg attcttcatt ggtcagaaaa ttatgaacgg
     4501 tttcttctat tttgtctcta tatactacgt ataggaaatg tttacatttt cgtattgttt
     4561 tcgattcact ctatgaatag ttcttactac aatttttttg tctaaagagt aatactagag
     4621 ataaacataa aaaatgtaga ggtcgagttt agatgcaagt tcaaggagcg aaaggtggat
     4681 gggtaggtta tatagggata tagcacagag atatatagca aagagatact tttgagcaat
     4741 gtttgtggaa gcggtattcg caatgggaag ctccaccccg gttgataatc agaaaagccc
     4801 caaaaacagg aagattgtat aagcaaatat ttaaattgta agcgttaata ttttgttaaa
     4861 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     4921 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     4981 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     5041 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     5101 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     5161 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     5221 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     5281 gggcgcgtgg ggatcgatcc actagtccac ggattagaag ccgccgagcg ggtgacagcc
     5341 ctccgaagga agactctcct ccgtgcgtcc tcgtcttcac cggtcgcgtt cctgaaacgc
     5401 agatgtgcct cgcgccgcac tgctccgaac aataaagatt ctacaatact agcttttatg
     5461 gttatgaaga ggaaaaattg gcagtaacct ggccccacaa accttcaaat gaacgaatca
     5521 aattaacaac cataggatga taatgcgatt agttttttag ccttatttct ggggtaatta
     5581 atcagcgaag cgatgatttt tgatctatta acagatatat aaatgcaaaa actgcataac
     5641 cactttaact aatactttca acattttcgg tttgtattac ttcttattca aatgtaataa
     5701 aagtatcaac aaaaaattgt taatatacct ctatacttta acgtcaagga gaaaaaaccc
     5761 cggatcggac tactagcagc tgtaatacga ctcactatag ggaatattaa gcttggatca
     5821 tgaaggtttt gattgtcttg ttggctatct tcgctgcttt gcccttggct ctagctcaac
     5881 cggttatttc tactaccgtc ggttccgctg cagaaggctc tttggacaag agagaagctc
     5941 ggccggacgt catgcccatc atgggctcct cggtgtacat cacggtggag ctggccattg
     6001 ctgtgctggc catcctgggc aatgtgctgg tgtgctgggc cgtgtggctc aacagcaacc
     6061 tgcagaacgt caccaactac tttgtggtgt cactggcggc ggccgacatc gcagtgggtg
     6121 tgctcgccat cccctttgcc atcaccatca gcaccgggtt ctgcgctgcc tgccacggct
     6181 gcctcttcat tgcctgcttc gtcctggtcc tcacgcagag ctccatcttc agtctcctgg
     6241 ccatcgccat tgaccgctac attgccatcc gcatcccgct ccggtacaat ggcttggtga
     6301 ccggcacgag ggctaagggc atcattgcca tctgctgggt gctgtcgttt gccatcggcc
     6361 tgactcccat gctaggttgg aacaactgcg gtcagccaaa ggagggcaag aaccactccc
     6421 agggctgcgg ggagggccaa gtggcctgtc tctttgagga tgtggtcccc atgaactaca
     6481 tggtgtactt caacttcttt gcctgtgtgc tggtgcccct gctgctcatg ctgggtgtct
     6541 atttgcggat cttcctggcg gcgcgacgac agctgaagca gatggagagc cagcctctgc
     6601 cgggggagcg ggcacggtcc acactgcaga aggaggtcca tgctgccaag tcactggcca
     6661 tcattgtggg gctctttgcc ctctgctggc tgcccctaca catcatcaac tgcttcactt
     6721 tcttctgccc cgactgcagc cacgcccctc tctggctcat gtacctggcc atcgtcctct
     6781 cccacaccaa ttcggttgtg aatcccttca tctacgccta ccgtatccgc gagttccgcc
     6841 agaccttccg caagatcatt cgcagccacg tcctgaggca gcaagaacct ttcaaggcag
     6901 ctggcaccag tgcccgggtc ttggcagctc atggcagtga cggagagcag gtcagcctcc
     6961 gtctcaacgg ccacccgcca ggagtgtggg ccaacggcag tgctccccac cctgagcgga
     7021 ggcccaatgg ctacgccctg gggctggtga gtggagggag tgcccaagag tcccagggga
     7081 acacgggcct cccagacgtg gagctcctta gccatgagct caagggagtg tgcccagagc
     7141 cccctggcct agatgacccc ctggcccagg atggagcagg agtgtccccg cggatggtga
     7201 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     7261 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
     7321 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
     7381 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg
     7441 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
     7501 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     7561 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     7621 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca
     7681 aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact
     7741 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     7801 gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     7861 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag cacgtgtcat
     7921 cacatcatca tcatcatcat catcatcatc attaataact taagagttga agacatgggt
     7981 atctattact gtacggctcg agcatgcatc tagagggccg catcatgtaa ttagttatgt
     8041 cacgcttaca ttcacgccct ccccccacat ccgctctaac cgaaaaggaa ggagttagac
     8101 aacctgaagt ctaggtccct atttattttt ttatagttat gttagtatta agaacgttat
     8161 ttatatttca aatttttctt ttttttctgt acagacgcgt gtacgcatgt aacattatac
     8221 tgaaaacctt gcttgagaag gttttgggac gctcgaaggc tttaatttgc aagctgcggc
     8281 cctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     8341 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     8401 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     8461 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     8521 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     8581 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     8641 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     8701 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     8761 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     8821 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     8881 caggattagc agagcgaggt atgtaggcag tgctacagag ttcttgaagt ggtggcctaa
     8941 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     9001 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     9061 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     9121 cttttctacg gggtctgacg ctcagtgg
//
