LOCUS       sequence-311882-        8932 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express Gid10 (YGR066C) in yeast two hybrid plasmid, pGADCg.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8932)
  AUTHORS   Melnykov A, Chen SJ, Varshavsky A
  TITLE     Gid10 as an alternative N-recognin of the Pro/N-degron pathway.
  JOURNAL   Proc Natl Acad Sci U S A. 2019 Aug 6;116(32):15914-15923. doi: 
            10.1073/pnas.1908304116. Epub 2019 Jul 23.
  PUBMED    31337681
REFERENCE   2  (bases 1 to 8932)
  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: 
            "10.1073/pnas.1908304116"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2019-08-6- 6"; volume: "116"; issue: "32"; pages: 
            "15914-15923"
COMMENT     Sequence Label: pCSJ392
FEATURES             Location/Qualifiers
     source          1..8932
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        633..1337
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             1383..1403
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    1422..1446
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2331..2396
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    complement(2398..2422)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2436..2777
                     /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      3222..3409
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(3277..3301)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(3526..4620)
                     /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(4621..5026)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(5068..5084)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5068..5084)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5081..5103)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5092..5108
                     /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(5116..5122,5123..5140,5141..5146))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5161..5182
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    5237..5270
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    5348..5381
                     /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(5450..5467)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5621..6209)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5701..5720)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6380..7171,7172..7240))
                     /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     7003..7022
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7241..7345)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7413..7431
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7469..7491)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7591..7610
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      7627..8791
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 tttgcggcgc cgaaaaaacg agtttacgca attgcacaat catgctgact ctgtggcgga
       61 cccgcgctct tgccggcccg gcgataacgc tgggcgtgag gctgtgcccg gcggagtttt
      121 ttgcgcctgc attttccaag gtttaccctg cgctaagggg cgagattgga gaagcaataa
      181 gaatgccggt tggggttgcg atgatgacga ccacgacaac tggtgtcatt atttaagttg
      241 ccgaaagaac ctgagtgcat ttgcaacatg agtatactag aagaatgagc caagacttgc
      301 gagacgcgag tttgccggtg gtgcgaacaa tagagcgacc atgaccttga aggtgagacg
      361 cgcataaccg ctagagtact ttgaagagga aacagcaata gggttgctac cagtataaat
      421 agacaggtac atacaacact ggaaatggtt gtctgtttga gtacgctttc aattcatttg
      481 ggtgtgcact ttattatgtt acaatatgga agggaacttt acacttctcc tatgcacata
      541 tattaattaa agtccaatgc tagtagagaa ggggggtaac acccctccgc gctcttttcc
      601 gatttttttc taaaccgtgg aatatttcgg atatcctttt gttgtttccg ggtgtacaat
      661 atggacttcc tcttttctgg caaccaaacc catacatcgg gattcctata ataccttcgt
      721 tggtctccct aacatgtagg tggcggaggg gagatataca atagaacaga taccagacaa
      781 gacataatgg gctaaacaag actacaccaa ttacactgcc tcattgatgg tggtacataa
      841 cgaactaata ctgtagccct agacttgata gccatcatca tatcgaagtt tcactaccct
      901 ttttccattt gccatctatt gaagtaataa taggcgcatg caacttcttt tctttttttt
      961 tcttttctct ctcccccgtt gttgtctcac catatccgca atgacaaaaa aatgatggaa
     1021 gacactaaag gaaaaaatta acgacaaaga cagcaccaac agatgtcgtt gttccagagc
     1081 tgatgagggg tatctcgaag cacacgaaac tttttccttc cttcattcac gcacactact
     1141 ctctaatgag caacggtata cggccttcct tccagttact tgaatttgaa ataaaaaaaa
     1201 gtttgctgtc ttgctatcaa gtataaatag acctgcaatt attaatcttt tgtttcctcg
     1261 tcattgttct cgttcccttt cttccttgtt tctttttctg cacaatattt caagctatac
     1321 caagcataca atcaactcca agctttgcaa agatggataa agcggaatta attcccgagc
     1381 ctccaaaaaa gaagagaaag gtcgaattgg gtaccgcatc aacaagtttg tacaaaaaag
     1441 caggcttcat gacatcactg aatatcatgg gaagaaagtt tattctagaa agggccaaaa
     1501 gaaatgacaa cattgaagaa atatatactt cggcctatgt ctctctaccc tcctcaactg
     1561 atacacgttt gcctcacttc aaagcaaaag aagaggactg tgatgtttac gaggaaggga
     1621 ccaatttagt gggtaagaat gctaaatata cctacagatc ccttggtaga catttagatt
     1681 ttttgaggcc aggattgaga tttggcggtt cgcaatcgtc aaagtacacg tattatacgg
     1741 tggaagtgaa gattgatacc gtaaacttgc cactttacaa ggattcgcgt tctttggacc
     1801 cgcatgtcac aggtactttc actattaaaa atttaacacc agtcctagat aaagttgtta
     1861 ccctttttga agggtatgtc atcaattata accaattccc gttatgttca ctgcattggc
     1921 ccgctgagga gacgctcgac ccttatatgg ctcagagaga aagtgattgc tctcattgga
     1981 agaggtttgg tcattttggt tctgataact ggtcgttaac ggaaagaaat tttggtcagt
     2041 ataatcatga aagtgcagaa tttatgaatc aacgatatat ttatcttaaa tggaaggaaa
     2101 ggttcctact tgatgatgag gaacaagaaa atctgatgtt ggacgataat catcatctgg
     2161 aaggcgcgtc atttgaggga ttttactatg tgtgtttaga ccagctaaca ggttcagtgg
     2221 aaggctacta ctatcatccc gcttgtgaat tgtttcaaaa actcgaactc gttccaacca
     2281 actgcgacgc attgaacacc tactcaagtg gatttgaaat tgcccccggg gattacaagg
     2341 atcatgatgg tgattacaag gatcatgaca ttgactataa ggatgatgat gacaaagacc
     2401 cagctttctt gtacaaagtg gttgatggta ccgccgccaa ttttaatcaa agtgggaata
     2461 ttgctgatag ctcattgtcc ttcactttca ctaacagtag caacggtccg aacctcataa
     2521 caactcaaac aaattctcaa gcgctttcac aaccaattgc ctcctctaac gttcatgata
     2581 acttcatgaa taatgaaatc acggctagta aaattgatga tggtaataat tcaaaaccac
     2641 tgtcacctgg ttggacggac caaactgcgt ataacgcgtt tggaatcact acagggatgt
     2701 ttaataccac tacaatggat gatgtatata actatctatt cgatgatgaa gataccccac
     2761 caaacccaaa aaaagagatc ctcgagtacg acgtaccaga ttacgcttag tgatcacgat
     2821 acgggatcca tcgagctcga gctgcagatg aatcgtagat actgaaaaac cccgcaagtt
     2881 cacttcaact gtgcatcgtg caccatctca atttctttca tttatacatc gttttgcctt
     2941 cttttatgta actatactcc tctaagtttc aatcttggcc atgtaacctc tgatctatag
     3001 aattttttaa atgactagaa ttaatgccca tctttttttt ggacctaaat tcttcatgaa
     3061 aatatattac gagggcttat tcagaagctt tggacttctt cgccagaggt ttggtcaagt
     3121 ctccaatcaa ggttgtcggc ttgtctacct tgccagaaat ttacgaaaag atggaaaagg
     3181 gtcaaatcgt tggtagatac gttgttgaca cttctaaata agcgaatttc ttatgattta
     3241 tgatttttat tattaaataa gttataaaaa aaataagtgt atacaaattt taaagtgact
     3301 cttaggtttt aaaacgaaaa ttcttattct tgagtaactc tttcctgtag gtcaggttgc
     3361 tttctcaggt atagcatgag gtcgctctta ttgaccacac ctctaccggc cggtcgaaat
     3421 tcccctaccc tatgaacata ttccattttg taatttcgtg tcgtttctat tatgaatttc
     3481 atttataaag tttatgtaca aatatcataa aaaaagagaa tctttttaag caaggatttt
     3541 cttaacttct tcggcgacag catcaccgac ttcggtggta ctgttggaac cacctaaatc
     3601 accagttctg atacctgcat ccaaaacctt tttaactgca tcttcaatgg ccttaccttc
     3661 ttcaggcaag ttcaatgaca atttcaacat cattgcagca gacaagatag tggcgatagg
     3721 gttgacctta ttctttggca aatctggagc agaaccgtgg catggttcgt acaaaccaaa
     3781 tgcggtgttc ttgtctggca aagaggccaa ggacgcagat ggcaacaaac ccaaggaacc
     3841 tgggataacg gaggcttcat cggagatgat atcaccaaac atgttgctgg tgattataat
     3901 accatttagg tgggttgggt tcttaactag gatcatggcg gcagaatcaa tcaattgatg
     3961 ttgaaccttc aatgtaggaa attcgttctt gatggtttcc tccacagttt ttctccataa
     4021 tcttgaagag gccaaaacat tagctttatc caaggaccaa ataggcaatg gtggctcatg
     4081 ttgtagggcc atgaaagcgg ccattcttgt gattctttgc acttctggaa cggtgtattg
     4141 ttcactatcc caagcgacac catcaccatc gtcttccttt ctcttaccaa agtaaatacc
     4201 tcccactaat tctctgacaa caacgaagtc agtaccttta gcaaattgtg gcttgattgg
     4261 agataagtct aaaagagagt cggatgcaaa gttacatggt cttaagttgg cgtacaattg
     4321 aagttcttta cggattttta gtaaaccttg ttcaggtcta acactacctg taccccattt
     4381 aggaccaccc acagcaccta acaaaacggc atcagccttc ttggaggctt ccagcgcctc
     4441 atctggaagt gggacacctg tagcttcgat agcagcacca ccaattaaat gattttcgaa
     4501 atcgaacttg acattggaac gaacatcaga aatagcttta agaaccttaa tggcttcggc
     4561 tgtgatttct tgaccaacgt ggtcacctgg caaaacgacg atcttcttag gggcagacat
     4621 tagaatggta tatccttgaa atatatatat atattgctga aatgtaaaag gtaagaaaag
     4681 ttagaaagta agacgattgc taaccaccta ttggaaaaaa caataggtcc ttaaataata
     4741 ttgtcaactt caagtattgt gatgcaagca tttagtcatg aacgcttctc tattctatat
     4801 gaaaagccgg ttccggcgct ctcacctttc ctttttctcc caatttttca gttgaaaaag
     4861 gtatatgcgt caggcgacct ctgaaattaa caaaaaattt ccagtcatcg aatttgattc
     4921 tgtgcgatag cgcccctgtg tgttctcgtt atgttgagga aaaaaataat ggttgctaag
     4981 agattcgaac tcttgcatct tacgatacct gagtattccc acagttgggg gatctcgact
     5041 ctagctagag gatcaattcg taatcatgtc atagctgttt cctgtgtgaa attgttatcc
     5101 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     5161 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     5221 cctgtcgtgc cagctgataa cttcgtataa tgtatgctat acgaagttat taggtctgaa
     5281 gaggagttta cgtccagcca agctagcttg gctgcaggtc gagcggccgc gatccggaac
     5341 ccttaatata acttcgtata atgtatgcta tacgaagtta tcagctgcat taatgaatcg
     5401 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     5461 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     5521 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     5581 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     5641 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     5701 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     5761 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     5821 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     5881 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     5941 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     6001 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     6061 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     6121 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     6181 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     6241 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     6301 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     6361 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     6421 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     6481 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     6541 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     6601 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     6661 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     6721 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     6781 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     6841 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     6901 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     6961 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     7021 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     7081 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     7141 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     7201 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     7261 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     7321 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag
     7381 aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
     7441 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
     7501 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     7561 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     7621 accataacgc atttaagcat aaacacgcac tatgccgttc ttctcatgta tatatatata
     7681 caggcaacac gcagatatag gtgcgacgtg aacagtgagc tgtatgtgcg cagctcgcgt
     7741 tgcattttcg gaagcgctcg ttttcggaaa cgctttgaag ttcctattcc gaagttccta
     7801 ttctctagct agaaagtata ggaacttcag agcgcttttg aaaaccaaaa gcgctctgaa
     7861 gacgcacttt caaaaaacca aaaacgcacc ggactgtaac gagctactaa aatattgcga
     7921 ataccgcttc cacaaacatt gctcaaaagt atctctttgc tatatatctc tgtgctatat
     7981 ccctatataa cctacccatc cacctttcgc tccttgaact tgcatctaaa ctcgacctct
     8041 acatttttta tgtttatctc tagtattact ctttagacaa aaaaattgta gtaagaacta
     8101 ttcatagagt gaatcgaaaa caatacgaaa atgtaaacat ttcctatacg tagtatatag
     8161 agacaaaata gaagaaaccg ttcataattt tctgaccaat gaagaatcat caacgctatc
     8221 actttctgtt cacaaagtat gcgcaatcca catcggtata gaatataatc ggggatgcct
     8281 ttatcttgaa aaaatgcacc cgcagcttcg ctagtaatca gtaaacgcgg gaagtggagt
     8341 caggcttttt ttatggaaga gaaaatagac accaaagtag ccttcttcta accttaacgg
     8401 acctacagtg caaaaagtta tcaagagact gcattataga gcgcacaaag gagaaaaaaa
     8461 gtaatctaag atgctttgtt agaaaaatag cgctctcggg atgcattttt gtagaacaaa
     8521 aaagaagtat agattctttg ttggtaaaat agcgctctcg cgttgcattt ctgttctgta
     8581 aaaatgcagc tcagattctt tgtttgaaaa attagcgctc tcgcgttgca tttttgtttt
     8641 acaaaaatga agcacagatt cttcgttggt aaaatagcgc tttcgcgttg catttctgtt
     8701 ctgtaaaaat gcagctcaga ttctttgttt gaaaaattag cgctctcgcg ttgcattttt
     8761 gttctacaaa atgaagcaca gatgcttcgt tgcttgcatg cctgcaggtc gagatccggg
     8821 atcgaagaaa tgatggtaaa tgaaatagga aatcaaggag catgaaggca aaagacaaat
     8881 ataagggtcg aacgaaaaat aaagtgaaaa gtgttgatat gatgtatttg gc
//
