LOCUS       sequence-387458-        8427 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of GAL4 transcriptional activation domain (AD)-AP-1 
            sigma-1A fusion protein in yeast (yeast two-hybrid assays).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8427)
  AUTHORS   Mattera R, De Pace R, Bonifacino JS
  TITLE     The adaptor protein chaperone AAGAB stabilizes AP-4 complex 
            subunits.
  JOURNAL   Mol Biol Cell. 2022 Oct 1;33(12):ar109. doi: 
            10.1091/mbc.E22-05-0177. Epub 2022 Aug 17.
  PUBMED    35976721
REFERENCE   2  (bases 1 to 8427)
  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: "Mol Biol 
            Cell."; date: "2022-10-1"; volume: "33(12):ar109. doi"; pages: " 
            10.1091/mbc.E22-05-0177. Epub 2022 Aug 17"
COMMENT     Sequence Label: pGADT7-AP-1 sigma-1A
FEATURES             Location/Qualifiers
     source          1..8427
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        771..1475
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             1521..1541
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1557..1898
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="activation domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 activation domain
                     /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     primer_bind     1904..1923
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1904..1922
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1929..1938
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1941..1967
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     terminator      2855..3042
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2910..2934)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(3159..4253)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIEATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(4254..4659)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(4701..4717)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4701..4717)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4714..4736)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4725..4741
                     /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(4749..4755,4756..4773,4774..4779))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4794..4815
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    4870..4903
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    4981..5014
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(5083..5100)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5254..5842)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5334..5353)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6013..6804,6805..6873))
                     /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     6636..6655
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6874..6978)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7046..7064
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7102..7124)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7224..7243
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      7260..8424
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 tgcatgcctg caggtcgaga tccgggatcg aagaaatgat ggtaaatgaa ataggaaatc
       61 aaggagcatg aaggcaaaag acaaatataa gggtcgaacg aaaaataaag tgaaaagtgt
      121 tgatatgatg tatttggctt tgcggcgccg aaaaaacgag tttacgcaat tgcacaatca
      181 tgctgactct gtggcggacc cgcgctcttg ccggcccggc gataacgctg ggcgtgaggc
      241 tgtgcccggc ggagtttttt gcgcctgcat tttccaaggt ttaccctgcg ctaaggggcg
      301 agattggaga agcaataaga atgccggttg gggttgcgat gatgacgacc acgacaactg
      361 gtgtcattat ttaagttgcc gaaagaacct gagtgcattt gcaacatgag tatactagaa
      421 gaatgagcca agacttgcga gacgcgagtt tgccggtggt gcgaacaata gagcgaccat
      481 gaccttgaag gtgagacgcg cataaccgct agagtacttt gaagaggaaa cagcaatagg
      541 gttgctacca gtataaatag acaggtacat acaacactgg aaatggttgt ctgtttgagt
      601 acgctttcaa ttcatttggg tgtgcacttt attatgttac aatatggaag ggaactttac
      661 acttctccta tgcacatata ttaattaaag tccaatgcta gtagagaagg ggggtaacac
      721 ccctccgcgc tcttttccga tttttttcta aaccgtggaa tatttcggat atccttttgt
      781 tgtttccggg tgtacaatat ggacttcctc ttttctggca accaaaccca tacatcggga
      841 ttcctataat accttcgttg gtctccctaa catgtaggtg gcggagggga gatatacaat
      901 agaacagata ccagacaaga cataatgggc taaacaagac tacaccaatt acactgcctc
      961 attgatggtg gtacataacg aactaatact gtagccctag acttgatagc catcatcata
     1021 tcgaagtttc actacccttt ttccatttgc catctattga agtaataata ggcgcatgca
     1081 acttcttttc tttttttttc ttttctctct cccccgttgt tgtctcacca tatccgcaat
     1141 gacaaaaaaa tgatggaaga cactaaagga aaaaattaac gacaaagaca gcaccaacag
     1201 atgtcgttgt tccagagctg atgaggggta tctcgaagca cacgaaactt tttccttcct
     1261 tcattcacgc acactactct ctaatgagca acggtatacg gccttccttc cagttacttg
     1321 aatttgaaat aaaaaaaagt ttgctgtctt gctatcaagt ataaatagac ctgcaattat
     1381 taatcttttg tttcctcgtc attgttctcg ttccctttct tccttgtttc tttttctgca
     1441 caatatttca agctatacca agcatacaat caactccaag ctttgcaaag atggataaag
     1501 cggaattaat tcccgagcct ccaaaaaaga agagaaaggt cgaattgggt accgccgcca
     1561 attttaatca aagtgggaat attgctgata gctcattgtc cttcactttc actaacagta
     1621 gcaacggtcc gaacctcata acaactcaaa caaattctca agcgctttca caaccaattg
     1681 cctcctctaa cgttcatgat aacttcatga ataatgaaat cacggctagt aaaattgatg
     1741 atggtaataa ttcaaaacca ctgtcacctg gttggacgga ccaaactgcg tataacgcgt
     1801 ttggaatcac tacagggatg tttaatacca ctacaatgga tgatgtatat aactatctat
     1861 tcgatgatga agatacccca ccaaacccaa aaaaagagat ctttaatacg actcactata
     1921 gggcgagcgc cgccatggag tacccatacg acgtaccaga ttacgctcat atggccatgg
     1981 aggccagtga attcatgatg cggttcatgc tattattcag ccggcaggga aaactgcggc
     2041 tgcaaaaatg gtacctggcc acttcggaca aggaacggaa gaagatggtg cgcgagctca
     2101 tgcaggttgt cctggctcga aagcccaaga tgtgcagctt cctggagtgg agggacctca
     2161 aagttgtcta taagagatat gccagcctct acttctgctg cgccatcgag ggccaagaca
     2221 atgagctcat cacactggag ctgatccacc gatacgtgga gctcttagac aaatactttg
     2281 gcagtgtgtg cgagctggac atcatcttca actttgagaa ggcctacttc atcctggatg
     2341 agtttttgat ggggggggat gtccaggaca cctccaagaa gagtgtgctg aaagccatcg
     2401 agcaggctga cctactgcaa gaggaggatg agtcgccacg gagtgtgctg gaggagatgg
     2461 gtttggcact cgagctgcag atgaatcgta gatactgaaa aaccccgcaa gttcacttca
     2521 actgtgcatc gtgcaccatc tcaatttctt tcatttatac atcgttttgc cttcttttat
     2581 gtaactatac tcctctaagt ttcaatcttg gccatgtaac ctctgatcta tagaattttt
     2641 taaatgacta gaattaatgc ccatcttttt tttggaccta aattcttcat gaaaatatat
     2701 tacgagggct tattcagaag ctttggactt cttcgccaga ggtttggtca agtctccaat
     2761 caaggttgtc ggcttgtcta ccttgccaga aatttacgaa aagatggaaa agggtcaaat
     2821 cgttggtaga tacgttgttg acacttctaa ataagcgaat ttcttatgat ttatgatttt
     2881 tattattaaa taagttataa aaaaaataag tgtatacaaa ttttaaagtg actcttaggt
     2941 tttaaaacga aaattcttat tcttgagtaa ctctttcctg taggtcaggt tgctttctca
     3001 ggtatagcat gaggtcgctc ttattgacca cacctctacc ggccggtcga aattccccta
     3061 ccctatgaac atattccatt ttgtaatttc gtgtcgtttc tattatgaat ttcatttata
     3121 aagtttatgt acaaatatca taaaaaaaga gaatcttttt aagcaaggat tttcttaact
     3181 tcttcggcga cagcatcacc gacttcggtg gtactgttgg aaccacctaa atcaccagtt
     3241 ctgatacctg catccaaaac ctttttaact gcatcttcaa tggccttacc ttcttcaggc
     3301 aagttcaatg acaatttcaa catcattgca gcagacaaga tagtggcgat agggttgacc
     3361 ttattctttg gcaaatctgg agcagaaccg tggcatggtt cgtacaaacc aaatgcggtg
     3421 ttcttgtctg gcaaagaggc caaggacgca gatggcaaca aacccaagga acctgggata
     3481 acggaggctt catcggagat gatatcacca aacatgttgc tggtgattat aataccattt
     3541 aggtgggttg ggttcttaac taggatcatg gcggcagaat caatcaattg atgttgaacc
     3601 ttcaatgtag gaaattcgtt cttgatggtt tcctccacag tttttctcca taatcttgaa
     3661 gaggccaaaa cattagcttt atccaaggac caaataggca atggtggctc atgttgtagg
     3721 gccatgaaag cggccattct tgtgattctt tgcacttctg gaacggtgta ttgttcacta
     3781 tcccaagcga caccatcacc atcgtcttcc tttctcttac caaagtaaat acctcccact
     3841 aattctctga caacaacgaa gtcagtacct ttagcaaatt gtggcttgat tggagataag
     3901 tctaaaagag agtcggatgc aaagttacat ggtcttaagt tggcgtacaa ttgaagttct
     3961 ttacggattt ttagtaaacc ttgttcaggt ctaacactac ctgtacccca tttaggacca
     4021 cccacagcac ctaacaaaac ggcatcagcc ttcttggagg cttccagcgc ctcatctgga
     4081 agtgggacac ctgtagcttc gatagcagca ccaccaatta aatgattttc gaaatcgaac
     4141 ttgacattgg aacgaacatc agaaatagct ttaagaacct taatggcttc ggctgtgatt
     4201 tcttgaccaa cgtggtcacc tggcaaaacg acgatcttct taggggcaga cattagaatg
     4261 gtatatcctt gaaatatata tatatattgc tgaaatgtaa aaggtaagaa aagttagaaa
     4321 gtaagacgat tgctaaccac ctattggaaa aaacaatagg tccttaaata atattgtcaa
     4381 cttcaagtat tgtgatgcaa gcatttagtc atgaacgctt ctctattcta tatgaaaagc
     4441 cggttccggc gctctcacct ttcctttttc tcccaatttt tcagttgaaa aaggtatatg
     4501 cgtcaggcga cctctgaaat taacaaaaaa tttccagtca tcgaatttga ttctgtgcga
     4561 tagcgcccct gtgtgttctc gttatgttga ggaaaaaaat aatggttgct aagagattcg
     4621 aactcttgca tcttacgata cctgagtatt cccacagttg ggggatctcg actctagcta
     4681 gaggatcaat tcgtaatcat gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     4741 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     4801 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     4861 tgccagctga taacttcgta taatgtatgc tatacgaagt tattaggtct gaagaggagt
     4921 ttacgtccag ccaagctagc ttggctgcag gtcgagcggc cgcgatccgg aacccttaat
     4981 ataacttcgt ataatgtatg ctatacgaag ttatcagctg cattaatgaa tcggccaacg
     5041 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     5101 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5161 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5221 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5281 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5341 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5401 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     5461 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     5521 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5581 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5641 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     5701 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5761 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5821 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5881 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5941 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6001 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6061 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6121 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6181 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6241 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6301 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6361 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6421 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6481 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6541 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6601 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6661 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6721 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6781 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6841 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6901 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6961 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     7021 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     7081 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     7141 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     7201 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccataa
     7261 cgcatttaag cataaacacg cactatgccg ttcttctcat gtatatatat atacaggcaa
     7321 cacgcagata taggtgcgac gtgaacagtg agctgtatgt gcgcagctcg cgttgcattt
     7381 tcggaagcgc tcgttttcgg aaacgctttg aagttcctat tccgaagttc ctattctcta
     7441 gctagaaagt ataggaactt cagagcgctt ttgaaaacca aaagcgctct gaagacgcac
     7501 tttcaaaaaa ccaaaaacgc accggactgt aacgagctac taaaatattg cgaataccgc
     7561 ttccacaaac attgctcaaa agtatctctt tgctatatat ctctgtgcta tatccctata
     7621 taacctaccc atccaccttt cgctccttga acttgcatct aaactcgacc tctacatttt
     7681 ttatgtttat ctctagtatt actctttaga caaaaaaatt gtagtaagaa ctattcatag
     7741 agtgaatcga aaacaatacg aaaatgtaaa catttcctat acgtagtata tagagacaaa
     7801 atagaagaaa ccgttcataa ttttctgacc aatgaagaat catcaacgct atcactttct
     7861 gttcacaaag tatgcgcaat ccacatcggt atagaatata atcggggatg cctttatctt
     7921 gaaaaaatgc acccgcagct tcgctagtaa tcagtaaacg cgggaagtgg agtcaggctt
     7981 tttttatgga agagaaaata gacaccaaag tagccttctt ctaaccttaa cggacctaca
     8041 gtgcaaaaag ttatcaagag actgcattat agagcgcaca aaggagaaaa aaagtaatct
     8101 aagatgcttt gttagaaaaa tagcgctctc gggatgcatt tttgtagaac aaaaaagaag
     8161 tatagattct ttgttggtaa aatagcgctc tcgcgttgca tttctgttct gtaaaaatgc
     8221 agctcagatt ctttgtttga aaaattagcg ctctcgcgtt gcatttttgt tttacaaaaa
     8281 tgaagcacag attcttcgtt ggtaaaatag cgctttcgcg ttgcatttct gttctgtaaa
     8341 aatgcagctc agattctttg tttgaaaaat tagcgctctc gcgttgcatt tttgttctac
     8401 aaaatgaagc acagatgctt cgttgct
//
