LOCUS       sequence-387834-        7100 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of GAL4 transcriptional activation domain (AD)-AP-4 
            sigma-4 fusion protein in yeast (yeast two-hybrid assays).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7100)
  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 7100)
  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: pGAD424-AP-4 sigma-4
FEATURES             Location/Qualifiers
     source          1..7100
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        5..406
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             452..472
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             488..829
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="activation domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 activation domain
                     /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     terminator      1678..1865
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1733..1757)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(1986..3080)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(3081..3486)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(3528..3544)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3528..3544)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3541..3563)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3552..3568
                     /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(3576..3582,3583..3600,3601..3606))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3621..3642
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3759..3776)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3930..4518)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4010..4029)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4689..5480,5481..5549))
                     /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     5312..5331
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5550..5654)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5722..5740
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5778..5800)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5900..5919
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      5936..7100
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 gcttgcatgc aacttctttt cttttttttt cttttctctc tcccccgttg ttgtctcacc
       61 atatccgcaa tgacaaaaaa atgatggaag acactaaagg aaaaaattaa cgacaaagac
      121 agcaccaaca gatgtcgttg ttccagagct gatgaggggt atctcgaagc acacgaaact
      181 ttttccttcc ttcattcacg cacactactc tctaatgagc aacggtatac ggccttcctt
      241 ccagttactt gaatttgaaa taaaaaaaag tttgctgtct tgctatcaag tataaataga
      301 cctgcaatta ttaatctttt gtttcctcgt cattgttctc gttccctttc ttccttgttt
      361 ctttttctgc acaatatttc aagctatacc aagcatacaa tcaactccaa gctttgcaaa
      421 gatggataaa gcggaattaa ttcccgagcc tccaaaaaag aagagaaagg tcgaattggg
      481 taccgccgcc aattttaatc aaagtgggaa tattgctgat agctcattgt ccttcacttt
      541 cactaacagt agcaacggtc cgaacctcat aacaactcaa acaaattctc aagcgctttc
      601 acaaccaatt gcctcctcta acgttcatga taacttcatg aataatgaaa tcacggctag
      661 taaaattgat gatggtaata attcaaaacc actgtcacct ggttggacgg accaaactgc
      721 gtataacgcg tttggaatca ctacagggat gtttaatacc actacaatgg atgatgtata
      781 taactatcta ttcgatgatg aagatacccc accaaaccca aaaaaagaga tcgaattccc
      841 ggggatccga atgatcaagt ttttccttat ggtgaacaag caagggcaga cccgactgtc
      901 taagtactac gagcatgtgg acattaataa acgtgcgctt ctggagactg aggtcagcaa
      961 gagttgcctg tctcggtcca gcgaacaatg ctcattcatt gaatacaagg attttaaact
     1021 gatctaccgg caatatgcag ctctctttgt tgtggttgga gttaatgaca ctgagaatga
     1081 gatggctatc tatgaattta tacacaattt tgtggaagtt ttagatgggt acttcagccg
     1141 agtgagtgaa ttagatataa tgtttaattt ggataaagtt cacatcattt tggatgagat
     1201 ggtgttaaat ggctgcattg tggaaactaa cagagccaga attcttgccc ctctgctgat
     1261 tcttgacaag ctgtcggaaa gctgactcga cctgcagaga tctatgaatc gtagatactg
     1321 aaaaaccccg caagttcact tcaactgtgc atcgtgcacc atctcaattt ctttcattta
     1381 tacatcgttt tgccttcttt tatgtaacta tactcctcta agtttcaatc ttggccatgt
     1441 aacctctgat ctatagaatt ttttaaatga ctagaattaa tgcccatctt ttttttggac
     1501 ctaaattctt catgaaaata tattacgagg gcttattcag aagctttgga cttcttcgcc
     1561 agaggtttgg tcaagtctcc aatcaaggtt gtcggcttgt ctaccttgcc agaaatttac
     1621 gaaaagatgg aaaagggtca aatcgttggt agatacgttg ttgacacttc taaataagcg
     1681 aatttcttat gatttatgat ttttattatt aaataagtta taaaaaaaat aagtgtatac
     1741 aaattttaaa gtgactctta ggttttaaaa cgaaaattct tattcttgag taactctttc
     1801 ctgtaggtca ggttgctttc tcaggtatag catgaggtcg ctcttattga ccacacctct
     1861 accggcatgc cggtcgaaat tcccctaccc tatgaacata ttccattttg taatttcgtg
     1921 tcgtttctat tatgaatttc atttataaag tttatgtaca aatatcataa aaaaagagaa
     1981 tctttttaag caaggatttt cttaacttct tcggcgacag catcaccgac ttcggtggta
     2041 ctgttggaac cacctaaatc accagttctg atacctgcat ccaaaacctt tttaactgca
     2101 tcttcaatgg ccttaccttc ttcaggcaag ttcaatgaca atttcaacat cattgcagca
     2161 gacaagatag tggcgatagg gttgacctta ttctttggca aatctggagc agaaccgtgg
     2221 catggttcgt acaaaccaaa tgcggtgttc ttgtctggca aagaggccaa ggacgcagat
     2281 ggcaacaaac ccaaggaacc tgggataacg gaggcttcat cggagatgat atcaccaaac
     2341 atgttgctgg tgattataat accatttagg tgggttgggt tcttaactag gatcatggcg
     2401 gcagaatcaa tcaattgatg ttgaaccttc aatgtaggaa attcgttctt gatggtttcc
     2461 tccacagttt ttctccataa tcttgaagag gccaaaacat tagctttatc caaggaccaa
     2521 ataggcaatg gtggctcatg ttgtagggcc atgaaagcgg ccattcttgt gattctttgc
     2581 acttctggaa cggtgtattg ttcactatcc caagcgacac catcaccatc gtcttccttt
     2641 ctcttaccaa agtaaatacc tcccactaat tctctgacaa caacgaagtc agtaccttta
     2701 gcaaattgtg gcttgattgg agataagtct aaaagagagt cggatgcaaa gttacatggt
     2761 cttaagttgg cgtacaattg aagttcttta cggattttta gtaaaccttg ttcaggtcta
     2821 acactacctg taccccattt aggaccaccc acagcaccta acaaaacggc atcagccttc
     2881 ttggaggctt ccagcgcctc atctggaagt ggaacacctg tagcatcgat agcagcacca
     2941 ccaattaaat gattttcgaa atcgaacttg acattggaac gaacatcaga aatagcttta
     3001 agaaccttaa tggcttcggc tgtgatttct tgaccaacgt ggtcacctgg caaaacgacg
     3061 atcttcttag gggcagacat tagaatggta tatccttgaa atatatatat atattgctga
     3121 aatgtaaaag gtaagaaaag ttagaaagta agacgattgc taaccaccta ttggaaaaaa
     3181 caataggtcc ttaaataata ttgtcaactt caagtattgt gatgcaagca tttagtcatg
     3241 aacgcttctc tattctatat gaaaagccgg ttccggcgct ctcacctttc ctttttctcc
     3301 caatttttca gttgaaaaag gtatatgcgt caggcgacct ctgaaattaa caaaaaattt
     3361 ccagtcatcg aatttgattc tgtgcgatag cgcccctgtg tgttctcgtt atgttgagga
     3421 aaaaaataat ggttgctaag agattcgaac tcttgcatct tacgatacct gagtattccc
     3481 acagttgggg gatctcgact ctagctagag gatcaattcg taatcatgtc atagctgttt
     3541 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     3601 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     3661 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     3721 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     3781 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     3841 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     3901 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     3961 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4021 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4081 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4141 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4201 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4261 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4321 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4381 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4441 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4501 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4561 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4621 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4681 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     4741 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     4801 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     4861 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     4921 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     4981 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     5041 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5101 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5161 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5221 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5281 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5341 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5401 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5461 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     5521 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     5581 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     5641 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
     5701 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc
     5761 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
     5821 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
     5881 cggggctggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccataacgca
     5941 tttaagcata aacacgcact atgccgttct tctcatgtat atatatatac aggcaacacg
     6001 cagatatagg tgcgacgtga acagtgagct gtatgtgcgc agctcgcgtt gcattttcgg
     6061 aagcgctcgt tttcggaaac gctttgaagt tcctattccg aagttcctat tctctagcta
     6121 gaaagtatag gaacttcaga gcgcttttga aaaccaaaag cgctctgaag acgcactttc
     6181 aaaaaaccaa aaacgcaccg gactgtaacg agctactaaa atattgcgaa taccgcttcc
     6241 acaaacattg ctcaaaagta tctctttgct atatatctct gtgctatatc cctatataac
     6301 ctacccatcc acctttcgct ccttgaactt gcatctaaac tcgacctcta cattttttat
     6361 gtttatctct agtattactc tttagacaaa aaaattgtag taagaactat tcatagagtg
     6421 aatcgaaaac aatacgaaaa tgtaaacatt tcctatacgt agtatataga gacaaaatag
     6481 aagaaaccgt tcataatttt ctgaccaatg aagaatcatc aacgctatca ctttctgttc
     6541 acaaagtatg cgcaatccac atcggtatag aatataatcg gggatgcctt tatcttgaaa
     6601 aaatgcaccc gcagcttcgc tagtaatcag taaacgcggg aagtggagtc aggctttttt
     6661 tatggaagag aaaatagaca ccaaagtagc cttcttctaa ccttaacgga cctacagtgc
     6721 aaaaagttat caagagactg cattatagag cgcacaaagg agaaaaaaag taatctaaga
     6781 tgctttgtta gaaaaatagc gctctcggga tgcatttttg tagaacaaaa aagaagtata
     6841 gattctttgt tggtaaaata gcgctctcgc gttgcatttc tgttctgtaa aaatgcagct
     6901 cagattcttt gtttgaaaaa ttagcgctct cgcgttgcat ttttgtttta caaaaatgaa
     6961 gcacagattc ttcgttggta aaatagcgct ttcgcgttgc atttctgttc tgtaaaaatg
     7021 cagctcagat tctttgtttg aaaaattagc gctctcgcgt tgcatttttg ttctacaaaa
     7081 tgaagcacag atgcttcgtt
//
