LOCUS       sequence-349660-        7596 bp    DNA     circular SYN 15-APR-2025
DEFINITION  pGBKT7 yeast two-hybrid plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7596)
  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 7596)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pTL322
FEATURES             Location/Qualifiers
     source          1..7596
                     /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      1631..1678
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1705..1892
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1760..1784)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(1916..1932)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1916..1932)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1929..1951)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1940..1956
                     /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(1964..1970,1971..1988,1989..1994))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2009..2030
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2147..2164)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2318..2906)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2398..2417)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3235..3282
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3307..3326
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(3514..4308)
                     /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(3514..4308)
                     /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(3625..3644)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4235..4254
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(4309..4413)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4440..5782
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(5403..5450)
                     /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     5823..5841
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5879..5901)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6001..6020
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        6041..6322
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             6323..6997
                     /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(7100..7555)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7237..7258)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7449..7468
                     /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 attcagcaag aaaactgtgc
     1321 tgcgcgtgga taccagctgc gacaaatgca agcgcaaaat cctgcagacc gtgagcggtc
     1381 tgcagggcgt ggacaagatc gatatcgaca gcgagaaagg caccatgacc gtgaccgcac
     1441 gcgcagatcc ggtggatgtg attgaacgca cccgcaaagc cggtaaacgc gccgaagtgg
     1501 tgaccattgg tccgccgccg gcaagcagca gcagtagcag cagcaatccg agcaccgagc
     1561 agcagcagat taaaaaacag gccccggccg ccgaagaaaa aggatccgtc gacctgcagc
     1621 ggccgcataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgcg
     1681 cgcttgcagc caagctaatt ccgggcgaat ttcttatgat ttatgatttt tattattaaa
     1741 taagttataa aaaaaataag tgtatacaaa ttttaaagtg actcttaggt tttaaaacga
     1801 aaattcttat tcttgagtaa ctctttcctg taggtcaggt tgctttctca ggtatagcat
     1861 gaggtcgctc ttattgacca cacctctacc ggcatgcaag cttggcgtaa tcatggtcat
     1921 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     1981 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     2041 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     2101 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     2161 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     2221 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     2281 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2341 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2401 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     2461 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     2521 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     2581 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     2641 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     2701 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     2761 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     2821 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     2881 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     2941 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     3001 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     3061 aacctgaggc tatggcaggg cctgccgccc cgacgttggc tgcgagccct gggccttcac
     3121 ccgaacttgg ggggtggggt ggggaaaagg aagaaacgcg ggcgtattgg ccccaatggg
     3181 gtctcggtgg ggtatcgaca gagtgccagc cctgggaccg aaccccgcgt ttatgaacaa
     3241 acgacccaac accgtgcgtt ttattctgtc tttttattgc cgtcatagcg cgggttcctt
     3301 ccggtattgt ctccttccgt gtttcagtta gcctccccct agggtgggcg aagaactcca
     3361 gcatgagatc cccgcgctgg aggatcatcc agccggcgtc ccggaaaacg attccgaagc
     3421 ccaacctttc atagaaggcg gcggtggaat cgaaatctcg tgatggcagg ttgggcgtcg
     3481 cttggtcggt catttcgaac cccagagtcc cgctcagaag aactcgtcaa gaaggcgata
     3541 gaaggcgatg cgctgcgaat cgggagcggc gataccgtaa agcacgagga agcggtcagc
     3601 ccattcgccg ccaagctctt cagcaatatc acgggtagcc aacgctatgt cctgatagcg
     3661 gtccgccaca cccagccggc cacagtcgat gaatccagaa aagcggccat tttccaccat
     3721 gatattcggc aagcaggcat cgccatgagt cacgacgaga tcctcgccgt cgggcatgct
     3781 cgccttgagc ctggcgaaca gttcggctgg cgcgagcccc tgatgctctt cgtccagatc
     3841 atcctgatcg acaagaccgg cttccatccg agtacgtgct cgctcgatgc gatgtttcgc
     3901 ttggtggtcg aatgggcagg tagccggatc aagcgtatgc agccgccgca ttgcatcagc
     3961 catgatggat actttctcgg caggagcaag gtgagatgac aggagatcct gccccggcac
     4021 ttcgcccaat agcagccagt cccttcccgc ttcagtgaca acgtcgagca cagctgcgca
     4081 aggaacgccc gtcgtggcca gccacgatag ccgcgctgcc tcgtcttgca gttcattcag
     4141 ggcaccggac aggtcggtct tgacaaaaag aaccgggcgc ccctgcgctg acagccggaa
     4201 cacggcggca tcagagcagc cgattgtctg ttgtgcccag tcatagccga atagcctctc
     4261 cacccaagcg gccggagaac ctgcgtgcaa tccatcttgt tcaatcatac tcttcctttt
     4321 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     4381 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     4441 acgaagcatc tgtgcttcat tttgtagaac aaaaatgcaa cgcgagagcg ctaatttttc
     4501 aaacaaagaa tctgagctgc atttttacag aacagaaatg caacgcgaaa gcgctatttt
     4561 accaacgaag aatctgtgct tcatttttgt aaaacaaaaa tgcaacgcga gagcgctaat
     4621 ttttcaaaca aagaatctga gctgcatttt tacagaacag aaatgcaacg cgagagcgct
     4681 attttaccaa caaagaatct atacttcttt tttgttctac aaaaatgcat cccgagagcg
     4741 ctatttttct aacaaagcat cttagattac tttttttctc ctttgtgcgc tctataatgc
     4801 agtctcttga taactttttg cactgtaggt ccgttaaggt tagaagaagg ctactttggt
     4861 gtctattttc tcttccataa aaaaagcctg actccacttc ccgcgtttac tgattactag
     4921 cgaagctgcg ggtgcatttt ttcaagataa aggcatcccc gattatattc tataccgatg
     4981 tggattgcgc atactttgtg aacagaaagt gatagcgttg atgattcttc attggtcaga
     5041 aaattatgaa cggtttcttc tattttgtct ctatatacta cgtataggaa atgtttacat
     5101 tttcgtattg ttttcgattc actctatgaa tagttcttac tacaattttt ttgtctaaag
     5161 agtaatacta gagataaaca taaaaaatgt agaggtcgag tttagatgca agttcaagga
     5221 gcgaaaggtg gatgggtagg ttatataggg atatagcaca gagatatata gcaaagagat
     5281 acttttgagc aatgtttgtg gaagcggtat tcgcaatatt ttagtagctc gttacagtcc
     5341 ggtgcgtttt tggttttttg aaagtgcgtc ttcagagcgc ttttggtttt caaaagcgct
     5401 ctgaagttcc tatactttct agagaatagg aacttcggaa taggaacttc aaagcgtttc
     5461 cgaaaacgag cgcttccgaa aatgcaacgc gagctgcgca catacagctc actgttcacg
     5521 tcgcacctat atctgcgtgt tgcctgtata tatatataca tgagaagaac ggcatagtgc
     5581 gtgtttatgc ttaaatgcgt acttatatgc gtctatttat gtaggatgaa aggtagtcta
     5641 gtacctcctg tgatattatc ccattccatg cggggtatcg tatgcttcct tcagcactac
     5701 cctttagctg ttctatatgc tgccactcct caattggatt agtctcatcc ttcaatgcta
     5761 tcatttcctt tgatattgga tcatactaag aaaccattat tatcatgaca ttaacctata
     5821 aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc
     5881 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     5941 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg
     6001 cggcatcaga gcagattgta ctgagagtgc accatagatc aacgacatta ctatatatat
     6061 aatataggaa gcatttaata gaacagcatc gtaatatatg tgtactttgc agttatgacg
     6121 ccagatggca gtagtggaag atattcttta ttgaaaaata gcttgtcacc ttacgtacaa
     6181 tcttgatccg gagcttttct ttttttgccg attaagaatt aattcggtcg aaaaaagaaa
     6241 aggagagggc caagagggag ggcattggtg actattgagc acgtgagtat acgtgattaa
     6301 gcacacaaag gcagcttgga gtatgtctgt tattaatttc acaggtagtt ctggtccatt
     6361 ggtgaaagtt tgcggcttgc agagcacaga ggccgcagaa tgtgctctag attccgatgc
     6421 tgacttgctg ggtattatat gtgtgcccaa tagaaagaga acaattgacc cggttattgc
     6481 aaggaaaatt tcaagtcttg taaaagcata taaaaatagt tcaggcactc cgaaatactt
     6541 ggttggcgtg tttcgtaatc aacctaagga ggatgttttg gctctggtca atgattacgg
     6601 cattgatatc gtccaactgc atggagatga gtcgtggcaa gaataccaag agttcctcgg
     6661 tttgccagtt attaaaagac tcgtatttcc aaaagactgc aacatactac tcagtgcagc
     6721 ttcacagaaa cctcattcgt ttattccctt gtttgattca gaagcaggtg ggacaggtga
     6781 acttttggat tggaactcga tttctgactg ggttggaagg caagagagcc ccgaaagctt
     6841 acattttatg ttagctggtg gactgacgcc agaaaatgtt ggtgatgcgc ttagattaaa
     6901 tggcgttatt ggtgttgatg taagcggagg tgtggagaca aatggtgtaa aagactctaa
     6961 caaaatagca aatttcgtca aaaatgctaa gaaataggtt attactgagt agtatttatt
     7021 taagtattgt ttgtgcactt gccgatctat gcggtgtgaa ataccgcaca gatgcgtaag
     7081 gagaaaatac cgcatcagga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     7141 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     7201 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     7261 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     7321 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     7381 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     7441 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     7501 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtccatt
     7561 cgccattcag gctgcgcaac tgttgggaag ggcgat
//
