LOCUS       sequence-177442-        5792 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 5792)
  TITLE     Rathjen lab plasmids
REFERENCE   2  (bases 1 to 5792)
  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-chCALEBlong.cyt (aa 505-594)
FEATURES             Location/Qualifiers
     source          1..5792
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      5..1169
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        1174..1575
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             1603..2043
                     /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      2359..2546
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2414..2438)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2599..3273)
                     /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(3274..3375)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(3389..3405)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3389..3405)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3402..3424)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3413..3429
                     /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(3437..3443,3444..3461,3462..3467))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3482..3503
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3620..3637)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3791..4379)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3871..3890)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4550..5341,5342..5410))
                     /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     5173..5192
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5411..5515)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5583..5601
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5639..5661)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5761..5780
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cataacgcat ttaagcataa acacgcacta tgccgttctt ctcatgtata tatatataca
       61 ggcaacacgc agatataggt gcgacgtgaa cagtgagctg tatgtgcgca gctcgcgttg
      121 cattttcgga agcgctcgtt ttcggaaacg ctttgaagtt cctattccga agttcctatt
      181 ctctagctag aaagtatagg aacttcagag cgcttttgaa aaccaaaagc gctctgaaga
      241 cgcactttca aaaaaccaaa aacgcaccgg actgtaacga gctactaaaa tattgcgaat
      301 accgcttcca caaacattgc tcaaaagtat ctctttgcta tatatctctg tgctatatcc
      361 ctatataacc tacccatcca cctttcgctc cttgaacttg catctaaact cgacctctac
      421 attttttatg tttatctcta gtattactct ttagacaaaa aaattgtagt aagaactatt
      481 catagagtga atcgaaaaca atacgaaaat gtaaacattt cctatacgta gtatatagag
      541 acaaaataga agaaaccgtt cataattttc tgaccaatga agaatcatca acgctatcac
      601 tttctgttca caaagtatgc gcaatccaca tcggtataga atataatcgg ggatgccttt
      661 atcttgaaaa aatgcacccg cagcttcgct agtaatcagt aaacgcggga agtggagtca
      721 ggcttttttt atggaagaga aaatagacac caaagtagcc ttcttctaac cttaacggac
      781 ctacagtgca aaaagttatc aagagactgc attatagagc gcacaaagga gaaaaaaagt
      841 aatctaagat gctttgttag aaaaatagcg ctctcgggat gcatttttgt agaacaaaaa
      901 agaagtatag attctttgtt ggtaaaatag cgctctcgcg ttgcatttct gttctgtaaa
      961 aatgcagctc agattctttg tttgaaaaat tagcgctctc gcgttgcatt tttgttttac
     1021 aaaaatgaag cacagattct tcgttggtaa aatagcgctt tcgcgttgca tttctgttct
     1081 gtaaaaatgc agctcagatt ctttgtttga aaaattagcg ctctcgcgtt gcatttttgt
     1141 tctacaaaat gaagcacaga tgcttcgttg cttgcatgca acttcttttc tttttttttc
     1201 ttttctctct cccccgttgt tgtctcacca tatccgcaat gacaaaaaaa tgatggaaga
     1261 cactaaagga aaaaattaac gacaaagaca gcaccaacag atgtcgttgt tccagagctg
     1321 atgaggggta tctcgaagca cacgaaactt tttccttcct tcattcacgc acactactct
     1381 ctaatgagca acggtatacg gccttccttc cagttacttg aatttgaaat aaaaaaaagt
     1441 ttgctgtctt gctatcaagt ataaatagac ctgcaattat taatcttttg tttcctcgtc
     1501 attgttctcg ttccctttct tccttgtttc tttttctgca caatatttca agctatacca
     1561 agcatacaat caactccaag cttgaagcaa gcctcctgaa agatgaagct actgtcttct
     1621 atcgaacaag catgcgatat ttgccgactt aaaaagctca agtgctccaa agaaaaaccg
     1681 aagtgcgcca agtgtctgaa gaacaactgg gagtgtcgct actctcccaa aaccaaaagg
     1741 tctccgctga ctagggcaca tctgacagaa gtggaatcaa ggctagaaag actggaacag
     1801 ctatttctac tgatttttcc tcgagaagac cttgacatga ttttgaaaat ggattcttta
     1861 caggatataa aagcattgtt aacaggatta tttgtacaag ataatgtgaa taaagatgcc
     1921 gtcacagata gattggcttc agtggagact gatatgcctc taacattgag acagcataga
     1981 ataagtgcga catcatcatc ggaagagagt agtaacaaag gtcaaagaca gttgactgta
     2041 tcgccggaat tcctctacct gctcaagacg gagaacagca aactgcgcaa aaccaaatac
     2101 cgcactccgt ccgagctgca caacgacaac ttctcgctgt ccaccatcgc cgagggctcc
     2161 cacccgaacg atgaccctgg cgcaccccac aagctgcagg acccgctgaa gcctggcctg
     2221 aaggacgagg aaccgctcag catcctcagc accgcgccag aggagggcag caagggcgaa
     2281 ccagggggct gcggggtgcc ctgcctgcac aacaacctgg ggtagggatc cgtcgacctg
     2341 cagccaagct aattccgggc gaatttctta tgatttatga tttttattat taaataagtt
     2401 ataaaaaaaa taagtgtata caaattttaa agtgactctt aggttttaaa acgaaaattc
     2461 ttattcttga gtaactcttt cctgtaggtc aggttgcttt ctcaggtata gcatgaggtc
     2521 gctcttattg accacacctc taccggcatg ccggcaagtg cacaaacaat acttaaataa
     2581 atactactca gtaataacct atttcttagc atttttgacg aaatttgcta ttttgttaga
     2641 gtcttttaca ccatttgtct ccacacctcc gcttacatca acaccaataa cgccatttaa
     2701 tctaagcgca tcaccaacat tttctggcgt cagtccacca gctaacataa aatgtaagct
     2761 ttcggggctc tcttgccttc caacccagtc agaaatcgag ttccaatcca aaagttcacc
     2821 tgtcccacct gcttctgaat caaacaaggg aataaacgaa tgaggtttct gtgaagctgc
     2881 actgagtagt atgttgcagt cttttggaaa tacgagtctt ttaataactg gcaaaccgag
     2941 gaactcttgg tattcttgcc acgactcatc tccatgcagt tggacgatat caatgccgta
     3001 atcattgacc agagccaaaa catcctcctt aggttgatta cgaaacacgc caaccaagta
     3061 tttcggagtg cctgaactat ttttatatgc ttttacaaga cttgaaattt tccttgcaat
     3121 aaccgggtca attgttctct ttctattggg cacacatata atacccagca agtcagcatc
     3181 ggaatctaga gcacattctg cggcctctgt gctctgcaag ccgcaaactt tcaccaatgg
     3241 accagaacta cctgtgaaat taataacaga catactccaa gctgcctttg tgtgcttaat
     3301 cacgtatact cacgtgctca atagtcacca atgccctccc tcttggccct ctccttttct
     3361 tttttcgacc gaattaattc gtaatcatgt catagctgtt tcctgtgtga aattgttatc
     3421 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     3481 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     3541 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     3601 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3661 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3721 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3781 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3841 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3901 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3961 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     4021 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4081 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4141 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4201 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     4261 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4321 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4381 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4441 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     4501 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     4561 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     4621 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4681 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4741 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4801 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4861 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4921 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4981 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     5041 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     5101 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     5161 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     5221 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     5281 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     5341 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     5401 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     5461 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     5521 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     5581 aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc
     5641 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     5701 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg
     5761 cggcatcaga gcagattgta ctgagagtgc ac
//
