LOCUS       sequence-360585-        6818 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast-two-hybrid BD vector with pZ promoter (activated by ZPM).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6818)
  AUTHORS   Feuer E, Zimran G, Shpilman M, Mosquna A
  TITLE     A Modified Yeast Two-Hybrid Platform Enables Dynamic Control of 
            Expression Intensities to Unmask Properties of Protein-Protein 
            Interactions.
  JOURNAL   ACS Synth Biol. 2022 Jul 27. doi: 10.1021/acssynbio.2c00192.
  PUBMED    35895499
REFERENCE   2  (bases 1 to 6818)
  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: "ACS Synth 
            Biol. 2022 Jul 27. doi: 10.1021/acssynbio.2c00192."
COMMENT     Sequence Label: pBD-pZ-FLAG
FEATURES             Location/Qualifiers
     source          1..6818
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             126..149
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(191..210)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(192..210)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      232..419
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(287..311)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(472..1146)
                     /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(1147..1248)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(1262..1278)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1262..1278)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1275..1297)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1286..1302
                     /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(1310..1316,1317..1334,1335..1340))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1355..1376
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        complement(1448..1552)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1579..2034
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1666..1685)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1876..1897
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2129..2146)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2300..2888)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2380..2399)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3345..3447
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             3448..4107
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPELRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3514..3533)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     4388..4406
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4444..4466)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4566..4585
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      4602..5766
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     primer_bind     6441..6464
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             join(6494..6818,1..116)
                     /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"
ORIGIN
        1 atagattggc ttcagtggag actgatatgc ctctaacatt gagacagcat agaataagtg
       61 cgacatcatc atcggaagag agtagtaaca aaggtcaaag acagttgact gtatcgccgg
      121 aatttgacta caaggatgat gacgataaag cggaattcgc ccgggcctcg agcccgggtc
      181 gactctagag ccctatagtg agtcgtatta ctgcagccaa gctaattccg ggcgaatttc
      241 ttatgattta tgatttttat tattaaataa gttataaaaa aaataagtgt atacaaattt
      301 taaagtgact cttaggtttt aaaacgaaaa ttcttattct tgagtaactc tttcctgtag
      361 gtcaggttgc tttctcaggt atagcatgag gtcgctctta ttgaccacac ctctaccggc
      421 atgccggcaa gtgcacaaac aatacttaaa taaatactac tcagtaataa cctatttctt
      481 agcatttttg acgaaatttg ctattttgtt agagtctttt acaccatttg tctccacacc
      541 tccgcttaca tcaacaccaa taacgccatt taatctaagc gcatcaccaa cattttctgg
      601 cgtcagtcca ccagctaaca taaaatgtaa gctttcgggg ctctcttgcc ttccaaccca
      661 gtcagaaatc gagttccaat ccaaaagttc acctgtccca cctgcttctg aatcaaacaa
      721 gggaataaac gaatgaggtt tctgtgaagc tgcactgagt agtatgttgc agtcttttgg
      781 aaatacgagt cttttaataa ctggcaaacc gaggaactct tggtattctt gccacgactc
      841 atctccatgc agttggacga tatcaatgcc gtaatcattg accagagcca aaacatcctc
      901 cttaggttga ttacgaaaca cgccaaccaa gtatttcgga gtgcctgaac tatttttata
      961 tgcttttaca agacttgaaa ttttccttgc aataaccggg tcaattgttc tctttctatt
     1021 gggcacacat ataataccca gcaagtcagc atcggaatct agagcacatt ctgcggcctc
     1081 tgtgctctgc aagccgcaaa ctttcaccaa tggaccagaa ctacctgtga aattaataac
     1141 agacatactc caagctgcct ttgtgtgctt aatcacgtat actcacgtgc tcaatagtca
     1201 ccaatgccct ccctcttggc cctctccttt tcttttttcg accgaattaa ttcgtaatca
     1261 tgtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     1321 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     1381 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg gttctttccg
     1441 cctcaggact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     1501 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     1561 cccgaaaagt gccacctgac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     1621 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     1681 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     1741 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     1801 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     1861 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     1921 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     1981 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttacgcgc
     2041 ctaaattatg cggccgcata taactgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     2101 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     2161 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     2221 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     2281 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     2341 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     2401 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     2461 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     2521 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     2581 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     2641 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     2701 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     2761 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     2821 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     2881 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     2941 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     3001 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     3061 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     3121 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     3181 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     3241 agccagccgc ctgtgacgga agatcacttc gcagaataaa taaatcctgg tgtccctgtt
     3301 gataccggga agccctgggc caacttttgg cgaaaatgag acgttgatcg gcacgtaaga
     3361 ggttccaact ttcaccataa tgaaataaga tcactaccgg gcgtattttt tgagttgtcg
     3421 agattttcag gagctaagga agctaaaatg gagaaaaaaa tcactggata taccaccgtt
     3481 gatatatccc aatggcatcg taaagaacat tttgaggcat ttcagtcagt tgctcaatgt
     3541 acctataacc agaccgttca gctggatatt acggcctttt taaagaccgt aaagaaaaat
     3601 aagcacaagt tttatccggc ctttattcac attcttgccc gcctgatgaa tgctcatccg
     3661 gaattacgta tggcaatgaa agacggtgag ctggtgatat gggatagtgt tcacccttgt
     3721 tacaccgttt tccatgagca aactgaaacg ttttcatcgc tctggagtga ataccacgac
     3781 gatttccggc agtttctaca catatattcg caagatgtgg cgtgttacgg tgaaaacctg
     3841 gcctatttcc ctaaagggtt tattgagaat atgtttttcg tctcagccaa tccctgggtg
     3901 agtttcacca gttttgattt aaacgtggcc aatatggaca acttcttcgc ccccgttttc
     3961 accatgggca aatattatac gcaaggcgac aaggtgctga tgccgctggc gattcaggtt
     4021 catcatgccg tttgtgatgg cttccatgtc ggcagaatgc ttaatgaatt acaacagtac
     4081 tgcgatgagt ggcagggcgg ggcgtaattt ttttaaggca gttattggtg cccttaaacg
     4141 cctggttgct acgcctgaat aagtgataat aagcggatga atggcagaaa ttcgaaagca
     4201 aattcgaccc ggtcgtcggt tcagggcagg gtcgttaaat agccgcttat gtctattgct
     4261 ggtttaccgg tttattgact accggaagca gtgtgaccgt gtgcttctca aatgcctgag
     4321 gccagtttgc tcaggctctc cccgtggagg ctaagaaacc attattatca tgacattaac
     4381 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     4441 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     4501 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     4561 ctatgcggca tcagagcaga ttgtactgag agtgcaccat aacgcattta agcataaaca
     4621 cgcactatgc cgttcttctc atgtatatat atatacaggc aacacgcaga tataggtgcg
     4681 acgtgaacag tgagctgtat gtgcgcagct cgcgttgcat tttcggaagc gctcgttttc
     4741 ggaaacgctt tgaagttcct attccgaagt tcctattctc tagctagaaa gtataggaac
     4801 ttcagagcgc ttttgaaaac caaaagcgct ctgaagacgc actttcaaaa aaccaaaaac
     4861 gcaccggact gtaacgagct actaaaatat tgcgaatacc gcttccacaa acattgctca
     4921 aaagtatctc tttgctatat atctctgtgc tatatcccta tataacctac ccatccacct
     4981 ttcgctcctt gaacttgcat ctaaactcga cctctacatt ttttatgttt atctctagta
     5041 ttactcttta gacaaaaaaa ttgtagtaag aactattcat agagtgaatc gaaaacaata
     5101 cgaaaatgta aacatttcct atacgtagta tatagagaca aaatagaaga aaccgttcat
     5161 aattttctga ccaatgaaga atcatcaacg ctatcacttt ctgttcacaa agtatgcgca
     5221 atccacatcg gtatagaata taatcgggga tgcctttatc ttgaaaaaat gcacccgcag
     5281 cttcgctagt aatcagtaaa cgcgggaagt ggagtcaggc tttttttatg gaagagaaaa
     5341 tagacaccaa agtagccttc ttctaacctt aacggaccta cagtgcaaaa agttatcaag
     5401 agactgcatt atagagcgca caaaggagaa aaaaagtaat ctaagatgct ttgttagaaa
     5461 aatagcgctc tcgggatgca tttttgtaga acaaaaaaga agtatagatt ctttgttggt
     5521 aaaatagcgc tctcgcgttg catttctgtt ctgtaaaaat gcagctcaga ttctttgttt
     5581 gaaaaattag cgctctcgcg ttgcattttt gttttacaaa aatgaagcac agattcttcg
     5641 ttggtaaaat agcgctttcg cgttgcattt ctgttctgta aaaatgcagc tcagattctt
     5701 tgtttgaaaa attagcgctc tcgcgttgca tttttgttct acaaaatgaa gcacagatgc
     5761 ttcgttgcgc ggtaccgggc ccttatattg aattttcaaa aattcttact ttttttttgg
     5821 atggacgcaa agaagtttaa taatcatatt acatggcatt accaccatat acatatccat
     5881 atacatatcc atatctaatc ttacttatat gttgtggaaa tgtaaagagc cccattatct
     5941 tagcctaaaa aaaccttctc tttggaactt tcagtaatac gcttaactgc tcattgctat
     6001 attgaagtgc ggccgcgtgg gcgtgcgtgg gcgggcgtgg gcgtgcgtgg gcgggcgtgg
     6061 gcgtgcgtgg gcgtctagac cgtgcgtcct cgtcttcacc ggtcgcgttc ctgaaacgca
     6121 gatgtgccta atgccgcact gctccgaaca ataaagattc tacaatacta gcttttatgg
     6181 ttatgaagag gaaaaattgg cagtaacctg gccccacaaa ccttcaaatt aacgaatcaa
     6241 attaacaacc ataggatgat aatgcgatta gttttttagc cttatttctg gggtaattaa
     6301 tcagcgaagc gatgattttt gatctattaa cagatatata aatggaaaag ctgcataacc
     6361 actttaacta atactttcaa cattttcagt ttgtattact tcttattcaa atgtcataaa
     6421 agtatcaaca aaaaattgtt aatatacctc tatactttaa cgtcaaggag aaaaaactat
     6481 actcgagaat aaaatgaagc tactgtcttc tatcgaacaa gcatgcgata tttgccgact
     6541 taaaaagctc aagtgctcca aagaaaaacc gaagtgcgcc aagtgtctga agaacaactg
     6601 ggagtgtcgc tactctccca aaaccaaaag gtctccgctg actagggcac atctgacaga
     6661 agtggaatca aggctagaaa gactggaaca gctatttcta ctgatttttc ctcgagaaga
     6721 ccttgacatg attttgaaaa tggattcttt acaggatata aaagcattgt taacaggatt
     6781 atttgtacaa gataatgtga ataaagatgc cgtcacag
//
