LOCUS       sequence-388877-        7521 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of 1) GAL4 DNA-binding domain (BD)-tyrosinase cytosolic 
            tail fusion protein, 2) HA epitope and nuclear localization signal 
            (NLS) AP-3 sigma-3A fusion protein in yeast (yeast three-hybrid 
            assays).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7521)
  AUTHORS   Mattera R, Boehm M, Chaudhuri R, Prabhu Y, Bonifacino JS
  TITLE     Conservation and diversification of dileucine signal recognition by 
            adaptor protein (AP) complex variants.
  JOURNAL   J Biol Chem. 2011 Jan 21;286(3):2022-30. doi: 
            10.1074/jbc.M110.197178. Epub 2010 Nov 19.
  PUBMED    21097499
REFERENCE   2  (bases 1 to 7521)
  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"; doi: 
            "10.1074/jbc.M110.197178"; journalName: "J Biol Chem"; date: 
            "2011-01-21- 21"; volume: "286"; issue: "3"; pages: "2022-30"
COMMENT     Sequence Label: pBridge-tyrosinase.sigma-3A
FEATURES             Location/Qualifiers
     source          1..7521
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(13..30)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(184..772)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(264..283)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(943..1734,1735..1803))
                     /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     1566..1585
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1804..1908)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1976..1994
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2032..2054)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2154..2173
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      2190..3354
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        3359..3760
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             3788..4228
                     /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      4696..4883
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(4751..4775)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(4936..5610)
                     /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(5611..5712)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(5726..5742)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5726..5742)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5739..5761)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5750..5766
                     /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(5774..5780,5781..5798,5799..5804))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5819..5840
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6034..6384
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /label=MET17 promoter
                     /note="expression is repressed by methionine"
     CDS             6406..6432
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             6412..6438
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             6454..6474
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      7173..7443
                     /gene="S. cerevisae PGK1"
                     /label=PGK1 terminator
                     /note="transcription terminator for 3-phosphoglycerate 
                     kinase gene"
ORIGIN
        1 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
       61 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
      121 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
      181 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
      241 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
      301 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
      361 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
      421 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
      481 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
      541 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
      601 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
      661 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
      721 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
      781 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
      841 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
      901 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
      961 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     1021 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     1081 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     1141 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     1201 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     1261 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     1321 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     1381 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     1441 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     1501 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     1561 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     1621 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     1681 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     1741 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     1801 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     1861 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     1921 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag
     1981 gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     2041 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     2101 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc
     2161 agagcagatt gtactgagag tgcaccataa cgcatttaag cataaacacg cactatgccg
     2221 ttcttctcat gtatatatat atacaggcaa cacgcagata taggtgcgac gtgaacagtg
     2281 agctgtatgt gcgcagctcg cgttgcattt tcggaagcgc tcgttttcgg aaacgctttg
     2341 aagttcctat tccgaagttc ctattctcta gctagaaagt ataggaactt cagagcgctt
     2401 ttgaaaacca aaagcgctct gaagacgcac tttcaaaaaa ccaaaaacgc accggactgt
     2461 aacgagctac taaaatattg cgaataccgc ttccacaaac attgctcaaa agtatctctt
     2521 tgctatatat ctctgtgcta tatccctata taacctaccc atccaccttt cgctccttga
     2581 acttgcatct aaactcgacc tctacatttt ttatgtttat ctctagtatt actctttaga
     2641 caaaaaaatt gtagtaagaa ctattcatag agtgaatcga aaacaatacg aaaatgtaaa
     2701 catttcctat acgtagtata tagagacaaa atagaagaaa ccgttcataa ttttctgacc
     2761 aatgaagaat catcaacgct atcactttct gttcacaaag tatgcgcaat ccacatcggt
     2821 atagaatata atcggggatg cctttatctt gaaaaaatgc acccgcagct tcgctagtaa
     2881 tcagtaaacg cgggaagtgg agtcaggctt tttttatgga agagaaaata gacaccaaag
     2941 tagccttctt ctaaccttaa cggacctaca gtgcaaaaag ttatcaagag actgcattat
     3001 agagcgcaca aaggagaaaa aaagtaatct aagatgcttt gttagaaaaa tagcgctctc
     3061 gggatgcatt tttgtagaac aaaaaagaag tatagattct ttgttggtaa aatagcgctc
     3121 tcgcgttgca tttctgttct gtaaaaatgc agctcagatt ctttgtttga aaaattagcg
     3181 ctctcgcgtt gcatttttgt tttacaaaaa tgaagcacag attcttcgtt ggtaaaatag
     3241 cgctttcgcg ttgcatttct gttctgtaaa aatgcagctc agattctttg tttgaaaaat
     3301 tagcgctctc gcgttgcatt tttgttctac aaaatgaagc acagatgctt cgttgcttgc
     3361 atgcaacttc ttttcttttt ttttcttttc tctctccccc gttgttgtct caccatatcc
     3421 gcaatgacaa aaaaatgatg gaagacacta aaggaaaaaa ttaacgacaa agacagcacc
     3481 aacagatgtc gttgttccag agctgatgag gggtatctcg aagcacacga aactttttcc
     3541 ttccttcatt cacgcacact actctctaat gagcaacggt atacggcctt ccttccagtt
     3601 acttgaattt gaaataaaaa aaagtttgct gtcttgctat caagtataaa tagacctgca
     3661 attattaatc ttttgtttcc tcgtcattgt tctcgttccc tttcttcctt gtttcttttt
     3721 ctgcacaata tttcaagcta taccaagcat acaatcaact ccaagcttga agcaagcctc
     3781 ctgaaagatg aagctactgt cttctatcga acaagcatgc gatatttgcc gacttaaaaa
     3841 gctcaagtgc tccaaagaaa aaccgaagtg cgccaagtgt ctgaagaaca actgggagtg
     3901 tcgctactct cccaaaacca aaaggtctcc gctgactagg gcacatctga cagaagtgga
     3961 atcaaggcta gaaagactgg aacagctatt tctactgatt tttcctcgag aagaccttga
     4021 catgattttg aaaatggatt ctttacagga tataaaagca ttgttaacag gattatttgt
     4081 acaagataat gtgaataaag atgccgtcac agatagattg gcttcagtgg agactgatat
     4141 gcctctaaca ttgagacagc atagaataag tgcgacatca tcatcggaag agagtagtaa
     4201 caaaggtcaa agacagttga ctgtatcgcc ggaattcccg gggatcctat gtcttcagaa
     4261 gaagaagaag aagaagcaac cccaggagga aaggcagcca ctcctcatgg acaaagacga
     4321 ctaccacagc ttgctgtatc agagccatct gtgaacatcc taggaaacag agtgggactg
     4381 aaaggtttta cctcactcga cctatttgtt ggtgtttcta caaatttaaa ctagtataaa
     4441 acatagacca tagctgtttg gctttttttc agacccatgt tttttcctaa gtcctagttt
     4501 ctaagaaatg actgggattt gctaaaatat atatatatat aaataataac ttactaatag
     4561 ctaaataaaa tttcctctta caactaattg agctggtttt tatgaatgtg tcttaattat
     4621 ttaaacttga ggcacatttt tgttttcctt acttcattgt gaatttccgg aattctgcag
     4681 ccaagctaat tccgggcgaa tttcttatga tttatgattt ttattattaa ataagttata
     4741 aaaaaaataa gtgtatacaa attttaaagt gactcttagg ttttaaaacg aaaattctta
     4801 ttcttgagta actctttcct gtaggtcagg ttgctttctc aggtatagca tgaggtcgct
     4861 cttattgacc acacctctac cggcatgccg gcaagtgcac aaacaatact taaataaata
     4921 ctactcagta ataacctatt tcttagcatt tttgacgaaa tttgctattt tgttagagtc
     4981 ttttacacca tttgtctcca cacctccgct tacatcaaca ccaataacgc catttaatct
     5041 aagcgcatca ccaacatttt ctggcgtcag tccaccagct aacataaaat gtaagctttc
     5101 ggggctctct tgccttccaa cccagtcaga aatcgagttc caatccaaaa gttcacctgt
     5161 cccacctgct tctgaatcaa acaagggaat aaacgaatga ggtttctgtg aagctgcact
     5221 gagtagtatg ttgcagtctt ttggaaatac gagtctttta ataactggca aaccgaggaa
     5281 ctcttggtat tcttgccacg actcatctcc atgcagttgg acgatatcaa tgccgtaatc
     5341 attgaccaga gccaaaacat cctccttagg ttgattacga aacacgccaa ccaagtattt
     5401 cggagtgcct gaactatttt tatatgcttt tacaagactt gaaattttcc ttgcaataac
     5461 cgggtcaatt gttctctttc tattgggcac acatataata cccagcaagt cagcatcgga
     5521 atctagagca cattctgcgg cctctgtgct ctgcaagccg caaactttca ccaatggacc
     5581 agaactacct gtgaaattaa taacagacat actccaagct gcctttgtgt gcttaatcac
     5641 gtatactcac gtgctcaata gtcaccaatg ccctccctct tggccctctc cttttctttt
     5701 ttcgaccgaa ttaattcgta atcatgtcat agctgtttcc tgtgtgaaat tgttatccgc
     5761 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     5821 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     5881 tgtcgtgcca ggaagatccg aggcctagct tctaattctt ccaacataca atgggagttt
     5941 ggccgagtgg tttaaggcgt cagatttagg tggatttaac ctctaaaatc tctgatatct
     6001 tcggatgcaa gggttcgaat cccttagctc tcattatttt ttgctttttc tcttgaggtc
     6061 acatgatcgc aaaatggcaa atggcacgtg aagctgtcga tattggggaa ctgtggtggt
     6121 tggcaaatga ctaattaagt tagtcaaggc gccatcctca tgaaaactgt gtaacataat
     6181 aaccgaagtg tcgaaaaggt ggcaccttgt ccaattgaac acgctcgatg aaaaaaataa
     6241 gatatatata aggttaagta aagcgtctgt tagaaaggaa gtttttcctt tttcttgctc
     6301 tcttgtcttt tcatctacta tttccttcgt gtaatacagg gtcgtcagat acatagatac
     6361 aattctatta cccccatcca tacaatgggc catatggctt ctagctatcc ttatgacgtg
     6421 cctgactatg ccagcctggg aggaccttct agtcctaaga agaagagaaa ggtggcggcc
     6481 gccatgatca aggcgatcct aatcttcaac aaccacggga agccgcggct ctccaagttc
     6541 taccagccct acagtgaaga tacacaacag caaatcatca gggagacttt ccatttggta
     6601 tctaagagag atgaaaatgt ttgtaatttc ctagaaggag gattattaat tggaggatct
     6661 gacaacaaac tgatttatag acattatgca acgttatatt ttgtcttctg tgtggattct
     6721 tcagaaagtg aacttggcat tttagatcta attcaagtat ttgtggaaac attagacaaa
     6781 tgttttgaaa atgtctgtga gctggatttg attttccatg tagacaaggt tcacaatatt
     6841 cttgcagaaa tggtgatggg gggaatggta ttggagacaa atatgaatga gattgttaca
     6901 caaattgatg cacaaaataa gctggaaaaa tctgaggctg gcttagcagg agctccagcc
     6961 cgtgctgtat cagctgtaaa gaatatgaat cttcctgaga tcccaagaaa tattaacatt
     7021 ggtgacatca gtataaaagt gccaaacctg ccctctttta aataaggatc ttcgggctga
     7081 tctcccatgt ctctactggt ggtggtgctt ctttggaatt attggaaggt aaggaattgc
     7141 caggtgttgc tttcttatcc gaaaagaaat aaattgaatt gaattgaaat cgatagatca
     7201 atttttttct tttctctttc cccatccttt acgctaaaat aatagtttat tttatttttt
     7261 gaatattttt tatttatata cgtatatata gactattatt tatcttttaa tgattattaa
     7321 gatttttatt aaaaaaaaat tcgctcctct tttaatgcct ttatgcagtt tttttttccc
     7381 attcgatatt tctatgttcg ggttcagcgt attttaagtt taataactcg aaaattctgc
     7441 gttcgttaaa gctaggcctc ggatcttcct gcattaatga atcggccaac gcgcggggag
     7501 aggcggtttg cgtattgggc g
//
