LOCUS       sequence-177451-        5667 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Two-hybrid system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5667)
  TITLE     Rathjen lab plasmids
REFERENCE   2  (bases 1 to 5667)
  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"
COMMENT     Sequence Label: pGBT9-chCALEBcyt (aa 505-551)
FEATURES             Location/Qualifiers
     source          1..5667
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..1165
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        1170..1571
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             1599..2039
                     /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"
     terminator      2230..2417
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2285..2309)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2470..3144)
                     /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"
     promoter        complement(3145..3246)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(3260..3276)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3260..3276)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3273..3295)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3284..3300
                     /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(3308..3314,3315..3332,3333..3338))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3353..3374
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3491..3508)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3662..4250)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3742..3761)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4421..5212,5213..5281))
                     /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     5044..5063
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5282..5386)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5454..5472
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5510..5532)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5632..5651
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 acgcatttaa gcataaacac gcactatgcc gttcttctca tgtatatata tatacaggca
       61 acacgcagat ataggtgcga cgtgaacagt gagctgtatg tgcgcagctc gcgttgcatt
      121 ttcggaagcg ctcgttttcg gaaacgcttt gaagttccta ttccgaagtt cctattctct
      181 agctagaaag tataggaact tcagagcgct tttgaaaacc aaaagcgctc tgaagacgca
      241 ctttcaaaaa accaaaaacg caccggactg taacgagcta ctaaaatatt gcgaataccg
      301 cttccacaaa cattgctcaa aagtatctct ttgctatata tctctgtgct atatccctat
      361 ataacctacc catccacctt tcgctccttg aacttgcatc taaactcgac ctctacattt
      421 tttatgttta tctctagtat tactctttag acaaaaaaat tgtagtaaga actattcata
      481 gagtgaatcg aaaacaatac gaaaatgtaa acatttccta tacgtagtat atagagacaa
      541 aatagaagaa accgttcata attttctgac caatgaagaa tcatcaacgc tatcactttc
      601 tgttcacaaa gtatgcgcaa tccacatcgg tatagaatat aatcggggat gcctttatct
      661 tgaaaaaatg cacccgcagc ttcgctagta atcagtaaac gcgggaagtg gagtcaggct
      721 ttttttatgg aagagaaaat agacaccaaa gtagccttct tctaacctta acggacctac
      781 agtgcaaaaa gttatcaaga gactgcatta tagagcgcac aaaggagaaa aaaagtaatc
      841 taagatgctt tgttagaaaa atagcgctct cgggatgcat ttttgtagaa caaaaaagaa
      901 gtatagattc tttgttggta aaatagcgct ctcgcgttgc atttctgttc tgtaaaaatg
      961 cagctcagat tctttgtttg aaaaattagc gctctcgcgt tgcatttttg ttttacaaaa
     1021 atgaagcaca gattcttcgt tggtaaaata gcgctttcgc gttgcatttc tgttctgtaa
     1081 aaatgcagct cagattcttt gtttgaaaaa ttagcgctct cgcgttgcat ttttgttcta
     1141 caaaatgaag cacagatgct tcgttgcttg catgcaactt cttttctttt tttttctttt
     1201 ctctctcccc cgttgttgtc tcaccatatc cgcaatgaca aaaaaatgat ggaagacact
     1261 aaaggaaaaa attaacgaca aagacagcac caacagatgt cgttgttcca gagctgatga
     1321 ggggtatctc gaagcacacg aaactttttc cttccttcat tcacgcacac tactctctaa
     1381 tgagcaacgg tatacggcct tccttccagt tacttgaatt tgaaataaaa aaaagtttgc
     1441 tgtcttgcta tcaagtataa atagacctgc aattattaat cttttgtttc ctcgtcattg
     1501 ttctcgttcc ctttcttcct tgtttctttt tctgcacaat atttcaagct ataccaagca
     1561 tacaatcaac tccaagcttg aagcaagcct cctgaaagat gaagctactg tcttctatcg
     1621 aacaagcatg cgatatttgc cgacttaaaa agctcaagtg ctccaaagaa aaaccgaagt
     1681 gcgccaagtg tctgaagaac aactgggagt gtcgctactc tcccaaaacc aaaaggtctc
     1741 cgctgactag ggcacatctg acagaagtgg aatcaaggct agaaagactg gaacagctat
     1801 ttctactgat ttttcctcga gaagaccttg acatgatttt gaaaatggat tctttacagg
     1861 atataaaagc attgttaaca ggattatttg tacaagataa tgtgaataaa gatgccgtca
     1921 cagatagatt ggcttcagtg gagactgata tgcctctaac attgagacag catagaataa
     1981 gtgcgacatc atcatcggaa gagagtagta acaaaggtca aagacagttg actgtatcgc
     2041 cggaattcct ctacctgctc aagacggaga acagcaaact gcgcaaaacc aaataccgca
     2101 ctccgtccga gctgcacaac gacaacttct cgctgtccac catcgccgag ggctcccacc
     2161 cgaacagaga agcgcagcat agagccctct aacctcggat ccgtcgacct gcagccaagc
     2221 taattccggg cgaatttctt atgatttatg atttttatta ttaaataagt tataaaaaaa
     2281 ataagtgtat acaaatttta aagtgactct taggttttaa aacgaaaatt cttattcttg
     2341 agtaactctt tcctgtaggt caggttgctt tctcaggtat agcatgaggt cgctcttatt
     2401 gaccacacct ctaccggcat gccggcaagt gcacaaacaa tacttaaata aatactactc
     2461 agtaataacc tatttcttag catttttgac gaaatttgct attttgttag agtcttttac
     2521 accatttgtc tccacacctc cgcttacatc aacaccaata acgccattta atctaagcgc
     2581 atcaccaaca ttttctggcg tcagtccacc agctaacata aaatgtaagc tttcggggct
     2641 ctcttgcctt ccaacccagt cagaaatcga gttccaatcc aaaagttcac ctgtcccacc
     2701 tgcttctgaa tcaaacaagg gaataaacga atgaggtttc tgtgaagctg cactgagtag
     2761 tatgttgcag tcttttggaa atacgagtct tttaataact ggcaaaccga ggaactcttg
     2821 gtattcttgc cacgactcat ctccatgcag ttggacgata tcaatgccgt aatcattgac
     2881 cagagccaaa acatcctcct taggttgatt acgaaacacg ccaaccaagt atttcggagt
     2941 gcctgaacta tttttatatg cttttacaag acttgaaatt ttccttgcaa taaccgggtc
     3001 aattgttctc tttctattgg gcacacatat aatacccagc aagtcagcat cggaatctag
     3061 agcacattct gcggcctctg tgctctgcaa gccgcaaact ttcaccaatg gaccagaact
     3121 acctgtgaaa ttaataacag acatactcca agctgccttt gtgtgcttaa tcacgtatac
     3181 tcacgtgctc aatagtcacc aatgccctcc ctcttggccc tctccttttc ttttttcgac
     3241 cgaattaatt cgtaatcatg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     3301 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     3361 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     3421 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     3481 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     3541 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     3601 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     3661 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     3721 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     3781 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     3841 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     3901 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     3961 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     4021 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     4081 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     4141 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     4201 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     4261 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     4321 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     4381 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     4441 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     4501 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     4561 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     4621 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     4681 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     4741 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     4801 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     4861 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     4921 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     4981 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     5041 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     5101 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     5161 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     5221 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     5281 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     5341 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     5401 aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc
     5461 gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
     5521 tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
     5581 gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag
     5641 agcagattgt actgagagtg caccata
//
