LOCUS       sequence-237452-        7642 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For Yeast-two-hybrid analysis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7642)
  AUTHORS   Liu X, Mao K, Yu AYH, Omairi-Nasser A, Austin J 2nd, Glick BS, Yip 
            CK, Klionsky DJ
  TITLE     The Atg17-Atg31-Atg29 Complex Coordinates with Atg11 to Recruit the 
            Vam7 SNARE and Mediate Autophagosome-Vacuole Fusion.
  JOURNAL   Curr Biol. 2016 Jan 25;26(2):150-160. doi: 
            10.1016/j.cub.2015.11.054. Epub 2016 Jan 7.
  PUBMED    26774783
REFERENCE   2  (bases 1 to 7642)
  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: 
            "10.1016/j.cub.2015.11.054"; journalName: "Curr Biol"; date: 
            "2016-01-25- 25"; volume: "26"; issue: "2"; pages: "150-160"
COMMENT     Sequence Label: pGAD-Atg17(∆335-417)
FEATURES             Location/Qualifiers
     source          1..7642
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        14..415
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             461..481
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             497..838
                     /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      2229..2416
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2284..2308)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2537..3631)
                     /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
                     LIDSAAMILVRNPTHLNGIIITSNMFGDIVSDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(3632..4037)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(4079..4095)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4079..4095)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4092..4114)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4103..4119
                     /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(4127..4133,4134..4151,4152..4157))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4172..4193
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4310..4327)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4481..5069)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4561..4580)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5240..6031,6032..6100))
                     /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     5863..5882
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6101..6205)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6273..6291
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6329..6351)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6451..6470
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      join(6487..7642,1..9)
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 tgcttcgttg cttgcatgca acttcttttc tttttttttc ttttctctct cccccgttgt
       61 tgtctcacca tatccgcaat gacaaaaaaa tgatggaaga cactaaagga aaaaattaac
      121 gacaaagaca gcaccaacag atgtcgttgt tccagagctg atgaggggta tctcgaagca
      181 cacgaaactt tttccttcct tcattcacgc acactactct ctaatgagca acggtatacg
      241 gccttccttc cagttacttg aatttgaaat aaaaaaaagt ttgctgtctt gctatcaagt
      301 ataaatagac ctgcaattat taatcttttg tttcctcgtc attgttctcg ttccctttct
      361 tccttgtttc tttttctgca caatatttca agctatacca agcatacaat caactccaag
      421 ctttgcaaag atggataaag cggaattaat tcccgagcct ccaaaaaaga agagaaaggt
      481 cgaattgggt accgccgcca attttaatca aagtgggaat attgctgata gctcattgtc
      541 cttcactttc actaacagta gcaacggtcc gaacctcata acaactcaaa caaattctca
      601 agcgctttca caaccaattg cctcctctaa cgttcatgat aacttcatga ataatgaaat
      661 cacggctagt aaaattgatg atggtaataa ttcaaaacca ctgtcacctg gttggacgga
      721 ccaaactgcg tataacgcgt ttggaatcac tacagggatg tttaatacca ctacaatgga
      781 tgatgtatat aactatctat tcgatgatga agatacccca ccaaacccaa aaaaagagat
      841 cgaattcccc gggggatcca tcgataacga agcagatgtt acaaaatttg ttaataatgc
      901 caggaaaacg ctgaccgatg ctcaactttt atgttcaagt gctaatttaa ggattgtaga
      961 tattaagaaa aaattgtcat cttggcagtt gagtatttca aaactcaatt ttctaatagt
     1021 tggcttgaga cagcaaggaa agtttcttta cactatttta aaggaaggca ttgggacaaa
     1081 gctaatccag aaacaatgga atcaagctgt cttagtggtc ttagttgatg agatgaagta
     1141 ctggcaatat gaaatcactt ctaaagtaca aaggttggat ggtatagtaa atgaattgag
     1201 catatcagaa aaagatgata ccgatccctc taaactagga gattacatct cgagagacaa
     1261 cgttaactta ttgaatgaca aactgaaaga agtgccagta attgagcgtc aaattgaaaa
     1321 cattaagctt caatacgaaa atatggtcag aaaagttaac aaagaactga ttgataccaa
     1381 gttgacggac gtaactcaaa aattccaaag taaatttggt atagataacc tgatggaaac
     1441 aaatgttgca gagcagttta gcagggaact aacggacctt gaaaaagatt tagcagagat
     1501 aatgaattca ttgacacagc actttgataa gacattgctg ttacaagata aaaaaattga
     1561 taacgatgaa cgtgaggagc tgtttaaggt ggtacaaggc gacgacaaag aactatacaa
     1621 cattttcaaa actctgcatg aggtaattga tgacgtggac aaaacaattc ttaacttggg
     1681 tcaatttttg caggcaaaaa taaaggaaaa gacagaacta cacagcgaag tttctgaaat
     1741 aataaacgat ttcaatagaa atttggaata tctattaatc tttaaagata tttccaatct
     1801 gattgatagc ttcaaaaatt cctgtacaca agatattcaa acaactaaag atctatgaat
     1861 cgtagatact gaaaaacccc gcaagttcac ttcaactgtg catcgtgcac catctcaatt
     1921 tctttcattt atacatcgtt ttgccttctt ttatgtaact atactcctct aagtttcaat
     1981 cttggccatg taacctctga tctatagaat tttttaaatg actagaatta atgcccatct
     2041 tttttttgga cctaaattct tcatgaaaat atattacgag ggcttattca gaagctttgg
     2101 acttcttcgc cagaggtttg gtcaagtctc caatcaaggt tgtcggcttg tctaccttgc
     2161 cagaaattta cgaaaagatg gaaaagggtc aaatcgttgg tagatacgtt gttgacactt
     2221 ctaaataagc gaatttctta tgatttatga tttttattat taaataagtt ataaaaaaaa
     2281 taagtgtata caaattttaa agtgactctt aggttttaaa acgaaaattc ttattcttga
     2341 gtaactcttt cctgtaggtc aggttgcttt ctcaggtata gcatgaggtc gctcttattg
     2401 accacacctc taccggcatg ccggtcgaaa ttcccctacc ctatgaacat attccatttt
     2461 gtaatttcgt gtcgtttcta ttatgaattt catttataaa gtttatgtac aaatatcata
     2521 aaaaaagaga atctttttaa gcaaggattt tcttaacctc ttcggcgaca gcatcaccga
     2581 cttcggtggt actgttggaa ccacctaaat caccagttct gatacctgca tccaaaacct
     2641 ttttaactgc atcttcaatg gccttacctt cttcaggcaa gttcaatgac aatttcaaca
     2701 tcattgcagc agacaagata gtggcgatag ggttgacctt attctttggc aaatctggag
     2761 cagaaccgtg gcatggttcg tacaaaccaa atgcggtgtt cttgtctggc aaagaggcca
     2821 aggacgcaga tggcaacaaa cccaaggaac ctgggataac ggaggcttca tcggagacga
     2881 tatcaccaaa catgttgctg gtgattataa taccatttag gtgggttggg ttcctaacta
     2941 ggatcatggc ggcagaatca atcaattgat gttgaacctt caatgtagga aattcgttct
     3001 tgatggtttc ctccacagtt tttctccata atcttgaaga ggccaaaaca ttagctttat
     3061 ccaaggacca aataggcaat ggtggctcat gttgtagggc catgaaagcg gccattcttg
     3121 tgattctttg cacttctgga acggtgtatt gttcactatc ccaagcgaca ccatcaccat
     3181 cgtcttcctt tctcttacca aagtaaatac ctcccactaa ttctctgaca acaacgaagt
     3241 cagtaccttt agcaaattgt ggcttgattg gagataagtc taaaagagag tcggatgcaa
     3301 agttacatgg tcttaagttg gcgtacaatt gaagttcttt acggattttt agtaaacctt
     3361 gttcaggtct aacactacct gtaccccatt taggaccacc cacagcacct aacaaaacgg
     3421 catcagcctt cttggaggct tccagcgcct catctggaag tgggacacct gtagcttcga
     3481 tagcagcacc accaattaaa tgattttcga aatcgaactt gacattggaa cgaacatcag
     3541 aaatagcttt aagaacctta atggcttcgg ctgtgatttc ttgaccaacg tggtcacctg
     3601 gcaaaacgac gatcttctta ggggcagaca ttagaatggt atatccttga aatatatata
     3661 tatattgctg aaatgtaaaa ggtaagaaaa gttagaaagt aagacgattg ctaaccacct
     3721 attggaaaaa acaataggtc cttaaataat attgtcaact tcaagtattg tgatgcaagc
     3781 atttagtcat gaacgcttct ctattctata tgaaaagccg gttccggcgc tctcaccttt
     3841 cctttttctc ccaatttttc agttgaaaaa ggtatatgcg tcaggcgacc tctgaaatta
     3901 acaaaaaatt tccagtcatc gaatttgatt ctgtgcgata gcgcccctgt gtgttctcgt
     3961 tatgttgagg aaaaaaataa tggttgctaa gagattcgaa ctcttgcatc ttacgatacc
     4021 tgagtattcc cacagttggg ggatctcgac tctagctaga ggatcaattc gtaatcatgt
     4081 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     4141 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     4201 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     4261 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     4321 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4381 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4441 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4501 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4561 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4621 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     4681 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     4741 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     4801 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     4861 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     4921 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     4981 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     5041 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     5101 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     5161 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     5221 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     5281 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     5341 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     5401 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     5461 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     5521 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     5581 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     5641 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     5701 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     5761 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     5821 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     5881 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     5941 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     6001 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     6061 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     6121 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     6181 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag
     6241 aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
     6301 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
     6361 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     6421 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     6481 accataacgc atttaagcat aaacacgcac tatgccgttc ttctcatgta tatatatata
     6541 caggcaacac gcagatatag gtgcgacgtg aacagtgagc tgtatgtgcg cagctcgcgt
     6601 tgcattttcg gaagcgctcg ttttcggaaa cgctttgaag ttcctattcc gaagttccta
     6661 ttctctagct agaaagtata ggaacttcag agcgcttttg aaaaccaaaa gcgctctgaa
     6721 gacgcacttt caaaaaacca aaaacgcacc ggactgtaac gagctactaa aatattgcga
     6781 ataccgcttc cacaaacatt gctcaaaagt atctctttgc tatatatctc tgtgctatat
     6841 ccctatataa cctacccatc cacctttcgc tccttgaact tgcatctaaa ctcgacctct
     6901 acatttttta tgtttatctc tagtattact ctttagacaa aaaaattgta gtaagaacta
     6961 ttcatagagt gaatcgaaaa caatacgaaa atgtaaacat ttcctatacg tagtatatag
     7021 agacaaaata gaagaaaccg ttcataattt tctgaccaat gaagaatcat caacgctatc
     7081 actttctgtt cacaaagtat gcgcaatcca catcggtata gaatataatc ggggatgcct
     7141 ttatcttgaa aaaatgcacc cgcagcttcg ctagtaatca gtaaacgcgg gaagtggagt
     7201 caggcttttt ttatggaaga gaaaatagac accaaagtag ccttcttcta accttaacgg
     7261 acctacagtg caaaaagtta tcaagagact gcattataga gcgcacaaag gagaaaaaaa
     7321 gtaatctaag atgctttgtt agaaaaatag cgctctcggg atgcattttt gtagaacaaa
     7381 aaagaagtat agattctttg ttggtaaaat agcgctctcg cgttgcattt ctgttctgta
     7441 aaaatgcagc tcagattctt tgtttgaaaa attagcgctc tcgcgttgca tttttgtttt
     7501 acaaaaatga agcacagatt cttcgttggt aaaatagcgc tttcgcgttg catttctgtt
     7561 ctgtaaaaat gcagctcaga ttctttgttt gaaaaattag cgctctcgcg ttgcattttt
     7621 gttctacaaa atgaagcaca ga
//
