LOCUS       sequence-177453-        5807 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 5807)
  TITLE     Rathjen lab plasmids
REFERENCE   2  (bases 1 to 5807)
  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-mCALEBcyt (aa 447-539)
FEATURES             Location/Qualifiers
     source          1..5807
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      11..1175
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        1180..1581
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             1609..2049
                     /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      2380..2567
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2435..2459)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2620..3294)
                     /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(3295..3396)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(3410..3426)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3410..3426)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3423..3445)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3434..3450
                     /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(3458..3464,3465..3482,3483..3488))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3503..3524
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3641..3658)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3812..4400)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3892..3911)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4571..5362,5363..5431))
                     /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     5194..5213
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5432..5536)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5604..5622
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5660..5682)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5782..5801
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gtgcaccata acgcatttaa gcataaacac gcactatgcc gttcttctca tgtatatata
       61 tatacaggca acacgcagat ataggtgcga cgtgaacagt gagctgtatg tgcgcagctc
      121 gcgttgcatt ttcggaagcg ctcgttttcg gaaacgcttt gaagttccta ttccgaagtt
      181 cctattctct agctagaaag tataggaact tcagagcgct tttgaaaacc aaaagcgctc
      241 tgaagacgca ctttcaaaaa accaaaaacg caccggactg taacgagcta ctaaaatatt
      301 gcgaataccg cttccacaaa cattgctcaa aagtatctct ttgctatata tctctgtgct
      361 atatccctat ataacctacc catccacctt tcgctccttg aacttgcatc taaactcgac
      421 ctctacattt tttatgttta tctctagtat tactctttag acaaaaaaat tgtagtaaga
      481 actattcata gagtgaatcg aaaacaatac gaaaatgtaa acatttccta tacgtagtat
      541 atagagacaa aatagaagaa accgttcata attttctgac caatgaagaa tcatcaacgc
      601 tatcactttc tgttcacaaa gtatgcgcaa tccacatcgg tatagaatat aatcggggat
      661 gcctttatct tgaaaaaatg cacccgcagc ttcgctagta atcagtaaac gcgggaagtg
      721 gagtcaggct ttttttatgg aagagaaaat agacaccaaa gtagccttct tctaacctta
      781 acggacctac agtgcaaaaa gttatcaaga gactgcatta tagagcgcac aaaggagaaa
      841 aaaagtaatc taagatgctt tgttagaaaa atagcgctct cgggatgcat ttttgtagaa
      901 caaaaaagaa gtatagattc tttgttggta aaatagcgct ctcgcgttgc atttctgttc
      961 tgtaaaaatg cagctcagat tctttgtttg aaaaattagc gctctcgcgt tgcatttttg
     1021 ttttacaaaa atgaagcaca gattcttcgt tggtaaaata gcgctttcgc gttgcatttc
     1081 tgttctgtaa aaatgcagct cagattcttt gtttgaaaaa ttagcgctct cgcgttgcat
     1141 ttttgttcta caaaatgaag cacagatgct tcgttgcttg catgcaactt cttttctttt
     1201 tttttctttt ctctctcccc cgttgttgtc tcaccatatc cgcaatgaca aaaaaatgat
     1261 ggaagacact aaaggaaaaa attaacgaca aagacagcac caacagatgt cgttgttcca
     1321 gagctgatga ggggtatctc gaagcacacg aaactttttc cttccttcat tcacgcacac
     1381 tactctctaa tgagcaacgg tatacggcct tccttccagt tacttgaatt tgaaataaaa
     1441 aaaagtttgc tgtcttgcta tcaagtataa atagacctgc aattattaat cttttgtttc
     1501 ctcgtcattg ttctcgttcc ctttcttcct tgtttctttt tctgcacaat atttcaagct
     1561 ataccaagca tacaatcaac tccaagcttg aagcaagcct cctgaaagat gaagctactg
     1621 tcttctatcg aacaagcatg cgatatttgc cgacttaaaa agctcaagtg ctccaaagaa
     1681 aaaccgaagt gcgccaagtg tctgaagaac aactgggagt gtcgctactc tcccaaaacc
     1741 aaaaggtctc cgctgactag ggcacatctg acagaagtgg aatcaaggct agaaagactg
     1801 gaacagctat ttctactgat ttttcctcga gaagaccttg acatgatttt gaaaatggat
     1861 tctttacagg atataaaagc attgttaaca ggattatttg tacaagataa tgtgaataaa
     1921 gatgccgtca cagatagatt ggcttcagtg gagactgata tgcctctaac attgagacag
     1981 catagaataa gtgcgacatc atcatcggaa gagagtagta acaaaggtca aagacagttg
     2041 actgtatcgc cggaattcct ctatctgctc aagactgaga ataccaagct gcggaggacc
     2101 aataaattcc ggaccccatc tgagctccac aacgacaact tctccctctc caccattgcc
     2161 gagggctctc atccaaatga cgaccccagc gctccccaca aaatccagga ccctctcaag
     2221 tcccgcctga aggaggaaga gtcctttaac atccagaact ccatgtcacc caaacttgag
     2281 ggtggcaaag gtgaccagga tgacttgggg gtgaactgtc tgcagaataa cctaacctga
     2341 gactgaggat ccgtcgacct gcagccaagc taattccggg cgaatttctt atgatttatg
     2401 atttttatta ttaaataagt tataaaaaaa ataagtgtat acaaatttta aagtgactct
     2461 taggttttaa aacgaaaatt cttattcttg agtaactctt tcctgtaggt caggttgctt
     2521 tctcaggtat agcatgaggt cgctcttatt gaccacacct ctaccggcat gccggcaagt
     2581 gcacaaacaa tacttaaata aatactactc agtaataacc tatttcttag catttttgac
     2641 gaaatttgct attttgttag agtcttttac accatttgtc tccacacctc cgcttacatc
     2701 aacaccaata acgccattta atctaagcgc atcaccaaca ttttctggcg tcagtccacc
     2761 agctaacata aaatgtaagc tttcggggct ctcttgcctt ccaacccagt cagaaatcga
     2821 gttccaatcc aaaagttcac ctgtcccacc tgcttctgaa tcaaacaagg gaataaacga
     2881 atgaggtttc tgtgaagctg cactgagtag tatgttgcag tcttttggaa atacgagtct
     2941 tttaataact ggcaaaccga ggaactcttg gtattcttgc cacgactcat ctccatgcag
     3001 ttggacgata tcaatgccgt aatcattgac cagagccaaa acatcctcct taggttgatt
     3061 acgaaacacg ccaaccaagt atttcggagt gcctgaacta tttttatatg cttttacaag
     3121 acttgaaatt ttccttgcaa taaccgggtc aattgttctc tttctattgg gcacacatat
     3181 aatacccagc aagtcagcat cggaatctag agcacattct gcggcctctg tgctctgcaa
     3241 gccgcaaact ttcaccaatg gaccagaact acctgtgaaa ttaataacag acatactcca
     3301 agctgccttt gtgtgcttaa tcacgtatac tcacgtgctc aatagtcacc aatgccctcc
     3361 ctcttggccc tctccttttc ttttttcgac cgaattaatt cgtaatcatg tcatagctgt
     3421 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     3481 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     3541 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     3601 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     3661 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     3721 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     3781 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     3841 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     3901 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     3961 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4021 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4081 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4141 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4201 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     4261 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     4321 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4381 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     4441 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     4501 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     4561 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     4621 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     4681 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     4741 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     4801 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     4861 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     4921 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     4981 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5041 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5101 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5161 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5221 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5281 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     5341 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     5401 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     5461 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     5521 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta
     5581 ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt
     5641 tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc
     5701 tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt
     5761 gtcggggctg gcttaactat gcggcatcag agcagattgt actgaga
//
