LOCUS       sequence-387734-        8424 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of GAL4 transcriptional activation domain (AD)-AP-1 
            sigma-1B fusion protein in yeast (yeast two-hybrid assays).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8424)
  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 8424)
  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-1B
FEATURES             Location/Qualifiers
     source          1..8424
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        749..1453
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             1499..1519
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1535..1876
                     /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     1882..1901
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1882..1900
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1907..1916
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1919..1945
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     terminator      2830..3017
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2885..2909)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(3134..4228)
                     /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(4229..4634)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(4676..4692)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4676..4692)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4689..4711)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4700..4716
                     /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(4724..4730,4731..4748,4749..4754))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4769..4790
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    4845..4878
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    4956..4989
                     /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(5058..5075)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5229..5817)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5309..5328)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5988..6779,6780..6848))
                     /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     6611..6630
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6849..6953)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7021..7039
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7077..7099)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7199..7218
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      7235..8399
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 cgggatcgaa gaaatgatgg taaatgaaat aggaaatcaa ggagcatgaa ggcaaaagac
       61 aaatataagg gtcgaacgaa aaataaagtg aaaagtgttg atatgatgta tttggctttg
      121 cggcgccgaa aaaacgagtt tacgcaattg cacaatcatg ctgactctgt ggcggacccg
      181 cgctcttgcc ggcccggcga taacgctggg cgtgaggctg tgcccggcgg agttttttgc
      241 gcctgcattt tccaaggttt accctgcgct aaggggcgag attggagaag caataagaat
      301 gccggttggg gttgcgatga tgacgaccac gacaactggt gtcattattt aagttgccga
      361 aagaacctga gtgcatttgc aacatgagta tactagaaga atgagccaag acttgcgaga
      421 cgcgagtttg ccggtggtgc gaacaataga gcgaccatga ccttgaaggt gagacgcgca
      481 taaccgctag agtactttga agaggaaaca gcaatagggt tgctaccagt ataaatagac
      541 aggtacatac aacactggaa atggttgtct gtttgagtac gctttcaatt catttgggtg
      601 tgcactttat tatgttacaa tatggaaggg aactttacac ttctcctatg cacatatatt
      661 aattaaagtc caatgctagt agagaagggg ggtaacaccc ctccgcgctc ttttccgatt
      721 tttttctaaa ccgtggaata tttcggatat ccttttgttg tttccgggtg tacaatatgg
      781 acttcctctt ttctggcaac caaacccata catcgggatt cctataatac cttcgttggt
      841 ctccctaaca tgtaggtggc ggaggggaga tatacaatag aacagatacc agacaagaca
      901 taatgggcta aacaagacta caccaattac actgcctcat tgatggtggt acataacgaa
      961 ctaatactgt agccctagac ttgatagcca tcatcatatc gaagtttcac tacccttttt
     1021 ccatttgcca tctattgaag taataatagg cgcatgcaac ttcttttctt tttttttctt
     1081 ttctctctcc cccgttgttg tctcaccata tccgcaatga caaaaaaatg atggaagaca
     1141 ctaaaggaaa aaattaacga caaagacagc accaacagat gtcgttgttc cagagctgat
     1201 gaggggtatc tcgaagcaca cgaaactttt tccttccttc attcacgcac actactctct
     1261 aatgagcaac ggtatacggc cttccttcca gttacttgaa tttgaaataa aaaaaagttt
     1321 gctgtcttgc tatcaagtat aaatagacct gcaattatta atcttttgtt tcctcgtcat
     1381 tgttctcgtt ccctttcttc cttgtttctt tttctgcaca atatttcaag ctataccaag
     1441 catacaatca actccaagct ttgcaaagat ggataaagcg gaattaattc ccgagcctcc
     1501 aaaaaagaag agaaaggtcg aattgggtac cgccgccaat tttaatcaaa gtgggaatat
     1561 tgctgatagc tcattgtcct tcactttcac taacagtagc aacggtccga acctcataac
     1621 aactcaaaca aattctcaag cgctttcaca accaattgcc tcctctaacg ttcatgataa
     1681 cttcatgaat aatgaaatca cggctagtaa aattgatgat ggtaataatt caaaaccact
     1741 gtcacctggt tggacggacc aaactgcgta taacgcgttt ggaatcacta cagggatgtt
     1801 taataccact acaatggatg atgtatataa ctatctattc gatgatgaag ataccccacc
     1861 aaacccaaaa aaagagatct ttaatacgac tcactatagg gcgagcgccg ccatggagta
     1921 cccatacgac gtaccagatt acgctcatat ggccatggag gccagtgaat tcatgcagtt
     1981 tatgttgctt tttagtcgtc agggaaagct tcgactgcaa aaatggtatg tcccactatc
     2041 agacaaagag aagaaaaaga tcacaagaga acttgttcag accgttttag cacggaaacc
     2101 taaaatgtgc agcttccttg agtggcgaga tctgaagatt gtttacaaaa gatatgctag
     2161 tctgtatttt tgctgtgcta ttgaggatca ggacaatgaa ctaattaccc tggaaataat
     2221 tcatcgttat gtggaattac ttgacaagta tttcggcagt gtctgtgaac tagatatcat
     2281 ctttaatttt gagaaggctt attttatttt ggatgagttt cttttgggag gggaagttca
     2341 ggaaacatcc aagaaaaatg tccttaaagc aattgagcag gctgatctac tgcaggagga
     2401 agctgaaacc ccacgtagtg ttcttgaaga aattggactg acactcgagc tgcagatgaa
     2461 tcgtagatac tgaaaaaccc cgcaagttca cttcaactgt gcatcgtgca ccatctcaat
     2521 ttctttcatt tatacatcgt tttgccttct tttatgtaac tatactcctc taagtttcaa
     2581 tcttggccat gtaacctctg atctatagaa ttttttaaat gactagaatt aatgcccatc
     2641 ttttttttgg acctaaattc ttcatgaaaa tatattacga gggcttattc agaagctttg
     2701 gacttcttcg ccagaggttt ggtcaagtct ccaatcaagg ttgtcggctt gtctaccttg
     2761 ccagaaattt acgaaaagat ggaaaagggt caaatcgttg gtagatacgt tgttgacact
     2821 tctaaataag cgaatttctt atgatttatg atttttatta ttaaataagt tataaaaaaa
     2881 ataagtgtat acaaatttta aagtgactct taggttttaa aacgaaaatt cttattcttg
     2941 agtaactctt tcctgtaggt caggttgctt tctcaggtat agcatgaggt cgctcttatt
     3001 gaccacacct ctaccggccg gtcgaaattc ccctacccta tgaacatatt ccattttgta
     3061 atttcgtgtc gtttctatta tgaatttcat ttataaagtt tatgtacaaa tatcataaaa
     3121 aaagagaatc tttttaagca aggattttct taacttcttc ggcgacagca tcaccgactt
     3181 cggtggtact gttggaacca cctaaatcac cagttctgat acctgcatcc aaaacctttt
     3241 taactgcatc ttcaatggcc ttaccttctt caggcaagtt caatgacaat ttcaacatca
     3301 ttgcagcaga caagatagtg gcgatagggt tgaccttatt ctttggcaaa tctggagcag
     3361 aaccgtggca tggttcgtac aaaccaaatg cggtgttctt gtctggcaaa gaggccaagg
     3421 acgcagatgg caacaaaccc aaggaacctg ggataacgga ggcttcatcg gagatgatat
     3481 caccaaacat gttgctggtg attataatac catttaggtg ggttgggttc ttaactagga
     3541 tcatggcggc agaatcaatc aattgatgtt gaaccttcaa tgtaggaaat tcgttcttga
     3601 tggtttcctc cacagttttt ctccataatc ttgaagaggc caaaacatta gctttatcca
     3661 aggaccaaat aggcaatggt ggctcatgtt gtagggccat gaaagcggcc attcttgtga
     3721 ttctttgcac ttctggaacg gtgtattgtt cactatccca agcgacacca tcaccatcgt
     3781 cttcctttct cttaccaaag taaatacctc ccactaattc tctgacaaca acgaagtcag
     3841 tacctttagc aaattgtggc ttgattggag ataagtctaa aagagagtcg gatgcaaagt
     3901 tacatggtct taagttggcg tacaattgaa gttctttacg gatttttagt aaaccttgtt
     3961 caggtctaac actacctgta ccccatttag gaccacccac agcacctaac aaaacggcat
     4021 cagccttctt ggaggcttcc agcgcctcat ctggaagtgg gacacctgta gcttcgatag
     4081 cagcaccacc aattaaatga ttttcgaaat cgaacttgac attggaacga acatcagaaa
     4141 tagctttaag aaccttaatg gcttcggctg tgatttcttg accaacgtgg tcacctggca
     4201 aaacgacgat cttcttaggg gcagacatta gaatggtata tccttgaaat atatatatat
     4261 attgctgaaa tgtaaaaggt aagaaaagtt agaaagtaag acgattgcta accacctatt
     4321 ggaaaaaaca ataggtcctt aaataatatt gtcaacttca agtattgtga tgcaagcatt
     4381 tagtcatgaa cgcttctcta ttctatatga aaagccggtt ccggcgctct cacctttcct
     4441 ttttctccca atttttcagt tgaaaaaggt atatgcgtca ggcgacctct gaaattaaca
     4501 aaaaatttcc agtcatcgaa tttgattctg tgcgatagcg cccctgtgtg ttctcgttat
     4561 gttgaggaaa aaaataatgg ttgctaagag attcgaactc ttgcatctta cgatacctga
     4621 gtattcccac agttggggga tctcgactct agctagagga tcaattcgta atcatgtcat
     4681 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     4741 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     4801 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgataact tcgtataatg
     4861 tatgctatac gaagttatta ggtctgaaga ggagtttacg tccagccaag ctagcttggc
     4921 tgcaggtcga gcggccgcga tccggaaccc ttaatataac ttcgtataat gtatgctata
     4981 cgaagttatc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     5041 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     5101 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     5161 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     5221 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     5281 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     5341 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     5401 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     5461 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     5521 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5581 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     5641 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     5701 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     5761 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     5821 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     5881 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     5941 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     6001 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     6061 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     6121 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     6181 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     6241 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     6301 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     6361 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     6421 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     6481 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     6541 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     6601 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     6661 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     6721 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     6781 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     6841 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     6901 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     6961 ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa
     7021 aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc
     7081 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     7141 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg
     7201 gcatcagagc agattgtact gagagtgcac cataacgcat ttaagcataa acacgcacta
     7261 tgccgttctt ctcatgtata tatatataca ggcaacacgc agatataggt gcgacgtgaa
     7321 cagtgagctg tatgtgcgca gctcgcgttg cattttcgga agcgctcgtt ttcggaaacg
     7381 ctttgaagtt cctattccga agttcctatt ctctagctag aaagtatagg aacttcagag
     7441 cgcttttgaa aaccaaaagc gctctgaaga cgcactttca aaaaaccaaa aacgcaccgg
     7501 actgtaacga gctactaaaa tattgcgaat accgcttcca caaacattgc tcaaaagtat
     7561 ctctttgcta tatatctctg tgctatatcc ctatataacc tacccatcca cctttcgctc
     7621 cttgaacttg catctaaact cgacctctac attttttatg tttatctcta gtattactct
     7681 ttagacaaaa aaattgtagt aagaactatt catagagtga atcgaaaaca atacgaaaat
     7741 gtaaacattt cctatacgta gtatatagag acaaaataga agaaaccgtt cataattttc
     7801 tgaccaatga agaatcatca acgctatcac tttctgttca caaagtatgc gcaatccaca
     7861 tcggtataga atataatcgg ggatgccttt atcttgaaaa aatgcacccg cagcttcgct
     7921 agtaatcagt aaacgcggga agtggagtca ggcttttttt atggaagaga aaatagacac
     7981 caaagtagcc ttcttctaac cttaacggac ctacagtgca aaaagttatc aagagactgc
     8041 attatagagc gcacaaagga gaaaaaaagt aatctaagat gctttgttag aaaaatagcg
     8101 ctctcgggat gcatttttgt agaacaaaaa agaagtatag attctttgtt ggtaaaatag
     8161 cgctctcgcg ttgcatttct gttctgtaaa aatgcagctc agattctttg tttgaaaaat
     8221 tagcgctctc gcgttgcatt tttgttttac aaaaatgaag cacagattct tcgttggtaa
     8281 aatagcgctt tcgcgttgca tttctgttct gtaaaaatgc agctcagatt ctttgtttga
     8341 aaaattagcg ctctcgcgtt gcatttttgt tctacaaaat gaagcacaga tgcttcgttg
     8401 cttgcatgcc tgcaggtcga gatc
//
