LOCUS       sequence-349014-        7542 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pGBKT7 yeast two-hybrid plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7542)
  AUTHORS   Bentham AR, Petit-Houdenot Y, Win J, Chuma I, Terauchi R, Banfield 
            MJ, Kamoun S, Langner T
  TITLE     A single amino acid polymorphism in a conserved effector of the 
            multihost blast fungus pathogen expands host-target binding 
            spectrum.
  JOURNAL   PLoS Pathog. 2021 Nov 10;17(11):e1009957. doi: 
            10.1371/journal.ppat.1009957. eCollection 2021 Nov.
  PUBMED    34758051
REFERENCE   2  (bases 1 to 7542)
  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: "PLoS 
            Pathog."; date: "2021-11-10"; volume: "17(11):e1009957. doi"; pages:
            " 10.1371/journal.ppat.1009957. eCollection 2021 Nov"
COMMENT     Sequence Label: pTL99
FEATURES             Location/Qualifiers
     source          1..7542
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        30..734
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             762..1202
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     primer_bind     1213..1232
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1213..1231
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1251..1280
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      1577..1624
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1651..1838
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1706..1730)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(1862..1878)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1862..1878)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1875..1897)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1886..1902
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1910..1916,1917..1934,1935..1940))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1955..1976
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2093..2110)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2264..2852)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2344..2363)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3181..3228
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3253..3272
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(3460..4254)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(3460..4254)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3571..3590)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4181..4200
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(4255..4359)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4386..5728
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(5349..5396)
                     /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     5769..5787
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5825..5847)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5947..5966
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5987..6268
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             6269..6943
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     rep_origin      complement(7046..7501)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7183..7204)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7395..7414
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cggtgcgggc ctcttcgcta ttacgccaga tccttttgtt gtttccgggt gtacaatatg
       61 gacttcctct tttctggcaa ccaaacccat acatcgggat tcctataata ccttcgttgg
      121 tctccctaac atgtaggtgg cggaggggag atatacaata gaacagatac cagacaagac
      181 ataatgggct aaacaagact acaccaatta cactgcctca ttgatggtgg tacataacga
      241 actaatactg tagccctaga cttgatagcc atcatcatat cgaagtttca ctaccctttt
      301 tccatttgcc atctattgaa gtaataatag gcgcatgcaa cttcttttct ttttttttct
      361 tttctctctc ccccgttgtt gtctcaccat atccgcaatg acaaaaaaat gatggaagac
      421 actaaaggaa aaaattaacg acaaagacag caccaacaga tgtcgttgtt ccagagctga
      481 tgaggggtat ctcgaagcac acgaaacttt ttccttcctt cattcacgca cactactctc
      541 taatgagcaa cggtatacgg ccttccttcc agttacttga atttgaaata aaaaaaagtt
      601 tgctgtcttg ctatcaagta taaatagacc tgcaattatt aatcttttgt ttcctcgtca
      661 ttgttctcgt tccctttctt ccttgtttct ttttctgcac aatatttcaa gctataccaa
      721 gcatacaatc aactccaagc ttgaagcaag cctcctgaaa gatgaagcta ctgtcttcta
      781 tcgaacaagc atgcgatatt tgccgactta aaaagctcaa gtgctccaaa gaaaaaccga
      841 agtgcgccaa gtgtctgaag aacaactggg agtgtcgcta ctctcccaaa accaaaaggt
      901 ctccgctgac tagggcacat ctgacagaag tggaatcaag gctagaaaga ctggaacagc
      961 tatttctact gatttttcct cgagaagacc ttgacatgat tttgaaaatg gattctttac
     1021 aggatataaa agcattgtta acaggattat ttgtacaaga taatgtgaat aaagatgccg
     1081 tcacagatag attggcttca gtggagactg atatgcctct aacattgaga cagcatagaa
     1141 taagtgcgac atcatcatcg gaagagagta gtaacaaagg tcaaagacag ttgactgtat
     1201 cgccggaatt tgtaatacga ctcactatag ggcgagccgc catcatggag gagcagaagc
     1261 tgatctcaga ggaggacctg catatggcca tggaggccga attcaaacag aaaattgtga
     1321 ttaaagtgca gatgacctgc gataaatgcc gtcgtagtgc cctgaccctg gccggtagca
     1381 cctatggtgt gcagagcgtg gccattgaag gtgaagaacg cgatcagctg gtggtggtgg
     1441 gtgatggtgt ggatgccacc agcctggcca gctgtctgcg caaagccgtg aaagtgggtc
     1501 gcgccgacat tatcaaagtg gaagccgtgg tggacgagaa aaaataagga tccgtcgacc
     1561 tgcagcggcc gcataactag cataacccct tggggcctct aaacgggtct tgaggggttt
     1621 tttgcgcgct tgcagccaag ctaattccgg gcgaatttct tatgatttat gatttttatt
     1681 attaaataag ttataaaaaa aataagtgta tacaaatttt aaagtgactc ttaggtttta
     1741 aaacgaaaat tcttattctt gagtaactct ttcctgtagg tcaggttgct ttctcaggta
     1801 tagcatgagg tcgctcttat tgaccacacc tctaccggca tgcaagcttg gcgtaatcat
     1861 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     1921 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     1981 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     2041 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     2101 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     2161 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     2221 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     2281 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     2341 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     2401 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     2461 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     2521 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2581 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2641 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2701 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2761 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2821 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2881 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2941 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     3001 atgagtaacc tgaggctatg gcagggcctg ccgccccgac gttggctgcg agccctgggc
     3061 cttcacccga acttgggggg tggggtgggg aaaaggaaga aacgcgggcg tattggcccc
     3121 aatggggtct cggtggggta tcgacagagt gccagccctg ggaccgaacc ccgcgtttat
     3181 gaacaaacga cccaacaccg tgcgttttat tctgtctttt tattgccgtc atagcgcggg
     3241 ttccttccgg tattgtctcc ttccgtgttt cagttagcct ccccctaggg tgggcgaaga
     3301 actccagcat gagatccccg cgctggagga tcatccagcc ggcgtcccgg aaaacgattc
     3361 cgaagcccaa cctttcatag aaggcggcgg tggaatcgaa atctcgtgat ggcaggttgg
     3421 gcgtcgcttg gtcggtcatt tcgaacccca gagtcccgct cagaagaact cgtcaagaag
     3481 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg
     3541 gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg
     3601 atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc
     3661 caccatgata ttcggcaagc aggcatcgcc atgagtcacg acgagatcct cgccgtcggg
     3721 catgctcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc
     3781 cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg
     3841 tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc
     3901 atcagccatg atggatactt tctcggcagg agcaaggtga gatgacagga gatcctgccc
     3961 cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt cgagcacagc
     4021 tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt cttgcagttc
     4081 attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct gcgctgacag
     4141 ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat agccgaatag
     4201 cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa tcatactctt
     4261 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     4321 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     4381 acctgaacga agcatctgtg cttcattttg tagaacaaaa atgcaacgcg agagcgctaa
     4441 tttttcaaac aaagaatctg agctgcattt ttacagaaca gaaatgcaac gcgaaagcgc
     4501 tattttacca acgaagaatc tgtgcttcat ttttgtaaaa caaaaatgca acgcgagagc
     4561 gctaattttt caaacaaaga atctgagctg catttttaca gaacagaaat gcaacgcgag
     4621 agcgctattt taccaacaaa gaatctatac ttcttttttg ttctacaaaa atgcatcccg
     4681 agagcgctat ttttctaaca aagcatctta gattactttt tttctccttt gtgcgctcta
     4741 taatgcagtc tcttgataac tttttgcact gtaggtccgt taaggttaga agaaggctac
     4801 tttggtgtct attttctctt ccataaaaaa agcctgactc cacttcccgc gtttactgat
     4861 tactagcgaa gctgcgggtg cattttttca agataaaggc atccccgatt atattctata
     4921 ccgatgtgga ttgcgcatac tttgtgaaca gaaagtgata gcgttgatga ttcttcattg
     4981 gtcagaaaat tatgaacggt ttcttctatt ttgtctctat atactacgta taggaaatgt
     5041 ttacattttc gtattgtttt cgattcactc tatgaatagt tcttactaca atttttttgt
     5101 ctaaagagta atactagaga taaacataaa aaatgtagag gtcgagttta gatgcaagtt
     5161 caaggagcga aaggtggatg ggtaggttat atagggatat agcacagaga tatatagcaa
     5221 agagatactt ttgagcaatg tttgtggaag cggtattcgc aatattttag tagctcgtta
     5281 cagtccggtg cgtttttggt tttttgaaag tgcgtcttca gagcgctttt ggttttcaaa
     5341 agcgctctga agttcctata ctttctagag aataggaact tcggaatagg aacttcaaag
     5401 cgtttccgaa aacgagcgct tccgaaaatg caacgcgagc tgcgcacata cagctcactg
     5461 ttcacgtcgc acctatatct gcgtgttgcc tgtatatata tatacatgag aagaacggca
     5521 tagtgcgtgt ttatgcttaa atgcgtactt atatgcgtct atttatgtag gatgaaaggt
     5581 agtctagtac ctcctgtgat attatcccat tccatgcggg gtatcgtatg cttccttcag
     5641 cactaccctt tagctgttct atatgctgcc actcctcaat tggattagtc tcatccttca
     5701 atgctatcat ttcctttgat attggatcat actaagaaac cattattatc atgacattaa
     5761 cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg
     5821 aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg
     5881 ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta
     5941 actatgcggc atcagagcag attgtactga gagtgcacca tagatcaacg acattactat
     6001 atatataata taggaagcat ttaatagaac agcatcgtaa tatatgtgta ctttgcagtt
     6061 atgacgccag atggcagtag tggaagatat tctttattga aaaatagctt gtcaccttac
     6121 gtacaatctt gatccggagc ttttcttttt ttgccgatta agaattaatt cggtcgaaaa
     6181 aagaaaagga gagggccaag agggagggca ttggtgacta ttgagcacgt gagtatacgt
     6241 gattaagcac acaaaggcag cttggagtat gtctgttatt aatttcacag gtagttctgg
     6301 tccattggtg aaagtttgcg gcttgcagag cacagaggcc gcagaatgtg ctctagattc
     6361 cgatgctgac ttgctgggta ttatatgtgt gcccaataga aagagaacaa ttgacccggt
     6421 tattgcaagg aaaatttcaa gtcttgtaaa agcatataaa aatagttcag gcactccgaa
     6481 atacttggtt ggcgtgtttc gtaatcaacc taaggaggat gttttggctc tggtcaatga
     6541 ttacggcatt gatatcgtcc aactgcatgg agatgagtcg tggcaagaat accaagagtt
     6601 cctcggtttg ccagttatta aaagactcgt atttccaaaa gactgcaaca tactactcag
     6661 tgcagcttca cagaaacctc attcgtttat tcccttgttt gattcagaag caggtgggac
     6721 aggtgaactt ttggattgga actcgatttc tgactgggtt ggaaggcaag agagccccga
     6781 aagcttacat tttatgttag ctggtggact gacgccagaa aatgttggtg atgcgcttag
     6841 attaaatggc gttattggtg ttgatgtaag cggaggtgtg gagacaaatg gtgtaaaaga
     6901 ctctaacaaa atagcaaatt tcgtcaaaaa tgctaagaaa taggttatta ctgagtagta
     6961 tttatttaag tattgtttgt gcacttgccg atctatgcgg tgtgaaatac cgcacagatg
     7021 cgtaaggaga aaataccgca tcaggaaatt gtaagcgtta atattttgtt aaaattcgcg
     7081 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     7141 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     7201 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     7261 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     7321 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     7381 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     7441 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     7501 tccattcgcc attcaggctg cgcaactgtt gggaagggcg at
//
