LOCUS       sequence-317847-        7610 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pYC2/CT 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 7610)
  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 7610)
  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: pCFEX2-A2aGH
FEATURES             Location/Qualifiers
     source          1..7610
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(32..823,824..892))
                     /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     655..674
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     RBS             2012..2020
                     /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)"
     misc_feature    2913..3417
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     3447..3466
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3575..4016
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    3575..3692
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3987..4010
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     4048..4067
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4048..4066
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(5459..5461,5462..5464,5465..6175)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(5459..5461,5462..5464,5465..6175)
                     /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(5504..5525)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5765..5784)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6112..6133
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             6188..6217
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             6286..6327
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             6337..6354
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      6388..6635
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(6404..6422)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(6712..6729)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6883..7471)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6963..6982)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
       61 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
      121 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
      181 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
      241 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
      301 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
      361 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
      421 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
      481 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
      541 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
      601 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
      661 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
      721 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
      781 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
      841 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactctact
      901 cctcgtcaat tctcttagga ttcgattcac attcatcttt ttttagctac taataggata
      961 aattatagga atttataaca aattgaaata tggcaggcag caaaattaaa attgtcttag
     1021 ttttttagtg tatagaagtg aatagctata taaagtatgt gtaaagttgg taacggaacg
     1081 aaaaatagaa aaggatatta catgggaaaa catgttgttt acggagaaat gaaaagtata
     1141 ttgtattttg tacgagctaa aagtacagtg ggaacaaagt cgattttgtt acatctacac
     1201 tgttgttatc agatctcgag ctattacaag tcttcttcag aaataagctt ttgttatcag
     1261 gagacggtga ctgaggttcc ttgaccccag tagtccatac catagtaaga acccgtacag
     1321 taatagatac ccatgtcttc aactcttaag ctaacaaatc gggttggtca atcctctagt
     1381 ccaagcatag gatccgagag ttatcaccaa taagcaatat gaaattgcaa tcgaaacagt
     1441 ataataaata agcatgttaa taaatgatag cttataccct tcattgataa acgtgctaat
     1501 ggtactcaaa gtgccgcaaa atccggatat taacgcagaa acaatatgac atgaattcaa
     1561 cgagttcaca attggaacgt ccgtagaaaa gtgtttttta ccacgttgca ccatggtaaa
     1621 tatgccaatc aataaagtgg cgaaaacgtt tgccaagaaa gtacccaatg gaaacttttt
     1681 gtttgtttta ttaaacattt ctgccaacca atatcttagg aaaccagcga aaattccaaa
     1741 tagacatggt agtgtccact tgcccctcga atagttctca taatacgcac acagcaccac
     1801 aaacaaaata attaatggaa aagctagtgc atacgcgagt ttatccacaa tatcaaaaaa
     1861 tttcttaaaa aatggtgctg gagttttgaa tttaaactct agaatatttt tttccatttc
     1921 ttgtgtatca acatcaccag cattttcctt tacagtgtct gagggagggg tatatggctt
     1981 cgaaaacgaa ctagatccat aagcaacaat aacctcctta ccaagttgcc taccgaaaat
     2041 aagactaccc atggaaacca taagatgaac gagcaagaca gataaaaact ccattatacc
     2101 gtaagctctg ttcggcagtt ttgtgtggtt agcgatatta ccgttagtta agtttgtaga
     2161 gtgttcgaac atctctagca gcatactaga aaaggaagac agggcaccac agtacccagt
     2221 agttacacct aaaaacaaga cttggtgatc tttcatccaa gtgtaagcat ttaaagattg
     2281 cataatcccc attagcatgc aggatgaaca attagaccag agaacggtcg gagcagtaac
     2341 gaaggccgga gtgtaggtgg aaagcgccgt tatagcctga cgtgtttcag tacctaatat
     2401 gcagaacgtg gtgaatgtac aaaaaacatg aatataatgg ctcaacttgt gatttgatat
     2461 gaccggattg aaaatcattt tgaattcttt ttaggctggt atcttgattc taaatcgatt
     2521 ttgcaagact agaggaatat gtattaaaat tcactgaatg caagtaaaat attaagtaaa
     2581 agaaagcatt gattatgaag aaaggacctt tttaagatat gagaacaatg agaaaaggaa
     2641 aaatgggacg aaacaaatag ggaaaattcc gtctgtaaaa ccaattgtag taacaaaaaa
     2701 cgctcatttc aatagcaaaa aaatagaata ccgtcaatgg atatatatag acacgggaca
     2761 ttataacaac gtgcagatgg tgatgattgc aaacccaaaa aattaccagc ttcctgtaat
     2821 tccaggttga gcgcgtggct tcctataatc caatacaccc acacccaccc agatcgtgat
     2881 ttttgttaca ggtcagctga tgggtccttt tcatcacgtg ctataaaaat aattataatt
     2941 taaatttttt aatataaata tataaattaa aaatagaaag taaaaaaaga aattaaagaa
     3001 aaaatagttt ttgttttccg aagatgtaaa agactctagg gggatcgcca acaaatacta
     3061 ccttttatct tgctcttcct gctctcaggt attaatgccg aattgtttca tcttgtctgt
     3121 gtagaagacc acacacgaaa atcctgtgat tttacatttt acttatcgtt aatcgaatgt
     3181 atatctattt aatctgcttt tcttgtctaa taaatatata tgtaaagtac gctttttgtt
     3241 gaaatttttt aaacctttgt ttattttttt tttcttcatt ccgtaactct tctaccttct
     3301 ttatttactt tctaaaatcc aaatacaaaa cataaaaata aataaacaca gagtaaattc
     3361 ccaaattatt ccatcattaa aagatacgag gcgcgtgtaa gttacaggca agcgatccgt
     3421 ccgccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     3481 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     3541 ctacagggcg cgtggggatc gatccactag tccacggatt agaagccgcc gagcgggtga
     3601 cagccctccg aaggaagact ctcctccgtg cgtcctcgtc ttcaccggtc gcgttcctga
     3661 aacgcagatg tgcctcgcgc cgcactgctc cgaacaataa agattctaca atactagctt
     3721 ttatggttat gaagaggaaa aattggcagt aacctggccc cacaaacctt caaatgaacg
     3781 aatcaaatta acaaccatag gatgataatg cgattagttt tttagcctta tttctggggt
     3841 aattaatcag cgaagcgatg atttttgatc tattaacaga tatataaatg caaaaactgc
     3901 ataaccactt taactaatac tttcaacatt ttcggtttgt attacttctt attcaaatgt
     3961 aataaaagta tcaacaaaaa attgttaata tacctctata ctttaacgtc aaggagaaaa
     4021 aaccccggat cggactacta gcagctgtaa tacgactcac tatagggaat attaagcttg
     4081 gatcatgaag gttttgattg tcttgttggc tatcttcgct gctttgccct tggctctagc
     4141 tcaaccggtt atttctacta ccgtcggttc cgctgcagaa ggctctttgg acaagagaga
     4201 agctcggccg gacgtcatgc ccatcatggg ctcctcggtg tacatcacgg tggagctggc
     4261 cattgctgtg ctggccatcc tgggcaatgt gctggtgtgc tgggccgtgt ggctcaacag
     4321 caacctgcag aacgtcacca actactttgt ggtgtcactg gcggcggccg acatcgcagt
     4381 gggtgtgctc gccatcccct ttgccatcac catcagcacc gggttctgcg ctgcctgcca
     4441 cggctgcctc ttcattgcct gcttcgtcct ggtcctcacg cagagctcca tcttcagtct
     4501 cctggccatc gccattgacc gctacattgc catccgcatc ccgctccggt acaatggctt
     4561 ggtgaccggc acgagggcta agggcatcat tgccatctgc tgggtgctgt cgtttgccat
     4621 cggcctgact cccatgctag gttggaacaa ctgcggtcag ccaaaggagg gcaagaacca
     4681 ctcccagggc tgcggggagg gccaagtggc ctgtctcttt gaggatgtgg tccccatgaa
     4741 ctacatggtg tacttcaact tctttgcctg tgtgctggtg cccctgctgc tcatgctggg
     4801 tgtctatttg cggatcttcc tggcggcgcg acgacagctg aagcagatgg agagccagcc
     4861 tctgccgggg gagcgggcac ggtccacact gcagaaggag gtccatgctg ccaagtcact
     4921 ggccatcatt gtggggctct ttgccctctg ctggctgccc ctacacatca tcaactgctt
     4981 cactttcttc tgccccgact gcagccacgc ccctctctgg ctcatgtacc tggccatcgt
     5041 cctctcccac accaattcgg ttgtgaatcc cttcatctac gcctaccgta tccgcgagtt
     5101 ccgccagacc ttccgcaaga tcattcgcag ccacgtcctg aggcagcaag aacctttcaa
     5161 ggcagctggc accagtgccc gggtcttggc agctcatggc agtgacggag agcaggtcag
     5221 cctccgtctc aacggccacc cgccaggagt gtgggccaac ggcagtgctc cccaccctga
     5281 gcggaggccc aatggctacg ccctggggct ggtgagtgga gggagtgccc aagagtccca
     5341 ggggaacacg ggcctcccag acgtggagct ccttagccat gagctcaagg gagtgtgccc
     5401 agagccccct ggcctagatg accccctggc ccaggatgga gcaggagtgt ccccgcggat
     5461 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     5521 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     5581 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     5641 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     5701 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     5761 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     5821 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     5881 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     5941 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     6001 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     6061 cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     6121 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagcacgt
     6181 gtcatcacat catcatcatc atcatcatca tcatcattaa taacttaaga gttgaagaca
     6241 tgggtatcta ttactgtacg gctcgagtct agagggccct tcgaaggtaa gcctatccct
     6301 aaccctctcc tcggtctcga ttctacgcgt accggtcatc atcaccatca ccattgagtt
     6361 taaacccgct gatcctagag ggccgcatca tgtaattagt tatgtcacgc ttacattcac
     6421 gccctccccc cacatccgct ctaaccgaaa aggaaggagt tagacaacct gaagtctagg
     6481 tccctattta tttttttata gttatgttag tattaagaac gttatttata tttcaaattt
     6541 ttcttttttt tctgtacaga cgcgtgtacg catgtaacat tatactgaaa accttgcttg
     6601 agaaggtttt gggacgctcg aaggctttaa tttgcaagct gcggccctgc attaatgaat
     6661 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     6721 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     6781 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     6841 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     6901 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     6961 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     7021 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     7081 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     7141 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     7201 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     7261 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     7321 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     7381 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     7441 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     7501 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     7561 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
//
