LOCUS       sequence-236138-        7792 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 7792)
  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 7792)
  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(∆266-312)
FEATURES             Location/Qualifiers
     source          1..7792
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             426..446
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             462..803
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="activation domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 activation domain
                     /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     RBS             1733..1744
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      2344..2531
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2399..2423)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2652..3746)
                     /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(3747..4152)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(4194..4210)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4194..4210)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4207..4229)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4218..4234
                     /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(4242..4248,4249..4266,4267..4272))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4287..4308
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4425..4442)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4596..5184)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4676..4695)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5355..6146,6147..6215))
                     /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     5978..5997
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6216..6320)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6388..6406
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6444..6466)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6566..6585
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      6602..7766
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        join(7771..7792,1..380)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
ORIGIN
        1 ttttcttttc tctctccccc gttgttgtct caccatatcc gcaatgacaa aaaaatgatg
       61 gaagacacta aaggaaaaaa ttaacgacaa agacagcacc aacagatgtc gttgttccag
      121 agctgatgag gggtatctcg aagcacacga aactttttcc ttccttcatt cacgcacact
      181 actctctaat gagcaacggt atacggcctt ccttccagtt acttgaattt gaaataaaaa
      241 aaagtttgct gtcttgctat caagtataaa tagacctgca attattaatc ttttgtttcc
      301 tcgtcattgt tctcgttccc tttcttcctt gtttcttttt ctgcacaata tttcaagcta
      361 taccaagcat acaatcaact ccaagctttg caaagatgga taaagcggaa ttaattcccg
      421 agcctccaaa aaagaagaga aaggtcgaat tgggtaccgc cgccaatttt aatcaaagtg
      481 ggaatattgc tgatagctca ttgtccttca ctttcactaa cagtagcaac ggtccgaacc
      541 tcataacaac tcaaacaaat tctcaagcgc tttcacaacc aattgcctcc tctaacgttc
      601 atgataactt catgaataat gaaatcacgg ctagtaaaat tgatgatggt aataattcaa
      661 aaccactgtc acctggttgg acggaccaaa ctgcgtataa cgcgtttgga atcactacag
      721 ggatgtttaa taccactaca atggatgatg tatataacta tctattcgat gatgaagata
      781 ccccaccaaa cccaaaaaaa gagatcgaat tccccggggg atccatcgat aacgaagcag
      841 atgttacaaa atttgttaat aatgccagga aaacgctgac cgatgctcaa cttttatgtt
      901 caagtgctaa tttaaggatt gtagatatta agaaaaaatt gtcatcttgg cagttgagta
      961 tttcaaaact caattttcta atagttggct tgagacagca aggaaagttt ctttacacta
     1021 ttttaaagga aggcattggg acaaagctaa tccagaaaca atggaatcaa gctgtcttag
     1081 tggtcttagt tgatgagatg aagtactggc aatatgaaat cacttctaaa gtacaaaggt
     1141 tggatggtat agtaaatgaa ttgagcatat cagaaaaaga tgataccgat ccctctaaac
     1201 taggagatta catctcgaga gacaacgtta acttattgaa tgacaaactg aaagaagtgc
     1261 cagtaattga gcgtcaaatt gaaaacatta agcttcaata cgaaaatatg gtcagaaaag
     1321 ttaacaaaga actgattgat accaagttga cggacgtaac tcaaaaattc caaagtaaat
     1381 ttggtataga taacctgatg gaaacaaatg ttgcagagca gtttagcagg gaactaacgg
     1441 accttgaaaa agatttagca gagataatga attcattgac acagcacttt gataagacat
     1501 tgctgttaca agataaaaaa attgataacg atgaacgtga ggagctgttt aaggtggtac
     1561 aaggcgacga caaagaacta tacaacattt tcaaaactct gcatgaggta attgatgacg
     1621 tgctgattga tagcttcaaa aattcctgta cacaagatat tcaaacaact aaggaacttt
     1681 gtgaatttta tgataatttt gaagaaagct acggtaactt agttctagaa gcaaagagga
     1741 gaaaggatgt ggcaaacaga atgaaaacta tattgaaaga ttgtgaaaag cagttgcaga
     1801 atttagatgc tcaggaccag gaagaacgtc agaattttat agcggaaaat ggaacttatc
     1861 ttcctgagac aatctggccc ggtaaaattg acgatttttc ttccctgtac actttaaatt
     1921 acaacgtgaa gaatccttag aaaaaaggaa aaaaaggctg cagagatcta tgaatcgtag
     1981 atactgaaaa accccgcaag ttcacttcaa ctgtgcatcg tgcaccatct caatttcttt
     2041 catttataca tcgttttgcc ttcttttatg taactatact cctctaagtt tcaatcttgg
     2101 ccatgtaacc tctgatctat agaatttttt aaatgactag aattaatgcc catctttttt
     2161 ttggacctaa attcttcatg aaaatatatt acgagggctt attcagaagc tttggacttc
     2221 ttcgccagag gtttggtcaa gtctccaatc aaggttgtcg gcttgtctac cttgccagaa
     2281 atttacgaaa agatggaaaa gggtcaaatc gttggtagat acgttgttga cacttctaaa
     2341 taagcgaatt tcttatgatt tatgattttt attattaaat aagttataaa aaaaataagt
     2401 gtatacaaat tttaaagtga ctcttaggtt ttaaaacgaa aattcttatt cttgagtaac
     2461 tctttcctgt aggtcaggtt gctttctcag gtatagcatg aggtcgctct tattgaccac
     2521 acctctaccg gcatgccggt cgaaattccc ctaccctatg aacatattcc attttgtaat
     2581 ttcgtgtcgt ttctattatg aatttcattt ataaagttta tgtacaaata tcataaaaaa
     2641 agagaatctt tttaagcaag gattttctta acctcttcgg cgacagcatc accgacttcg
     2701 gtggtactgt tggaaccacc taaatcacca gttctgatac ctgcatccaa aaccttttta
     2761 actgcatctt caatggcctt accttcttca ggcaagttca atgacaattt caacatcatt
     2821 gcagcagaca agatagtggc gatagggttg accttattct ttggcaaatc tggagcagaa
     2881 ccgtggcatg gttcgtacaa accaaatgcg gtgttcttgt ctggcaaaga ggccaaggac
     2941 gcagatggca acaaacccaa ggaacctggg ataacggagg cttcatcgga gacgatatca
     3001 ccaaacatgt tgctggtgat tataatacca tttaggtggg ttgggttcct aactaggatc
     3061 atggcggcag aatcaatcaa ttgatgttga accttcaatg taggaaattc gttcttgatg
     3121 gtttcctcca cagtttttct ccataatctt gaagaggcca aaacattagc tttatccaag
     3181 gaccaaatag gcaatggtgg ctcatgttgt agggccatga aagcggccat tcttgtgatt
     3241 ctttgcactt ctggaacggt gtattgttca ctatcccaag cgacaccatc accatcgtct
     3301 tcctttctct taccaaagta aatacctccc actaattctc tgacaacaac gaagtcagta
     3361 cctttagcaa attgtggctt gattggagat aagtctaaaa gagagtcgga tgcaaagtta
     3421 catggtctta agttggcgta caattgaagt tctttacgga tttttagtaa accttgttca
     3481 ggtctaacac tacctgtacc ccatttagga ccacccacag cacctaacaa aacggcatca
     3541 gccttcttgg aggcttccag cgcctcatct ggaagtggga cacctgtagc ttcgatagca
     3601 gcaccaccaa ttaaatgatt ttcgaaatcg aacttgacat tggaacgaac atcagaaata
     3661 gctttaagaa ccttaatggc ttcggctgtg atttcttgac caacgtggtc acctggcaaa
     3721 acgacgatct tcttaggggc agacattaga atggtatatc cttgaaatat atatatatat
     3781 tgctgaaatg taaaaggtaa gaaaagttag aaagtaagac gattgctaac cacctattgg
     3841 aaaaaacaat aggtccttaa ataatattgt caacttcaag tattgtgatg caagcattta
     3901 gtcatgaacg cttctctatt ctatatgaaa agccggttcc ggcgctctca cctttccttt
     3961 ttctcccaat ttttcagttg aaaaaggtat atgcgtcagg cgacctctga aattaacaaa
     4021 aaatttccag tcatcgaatt tgattctgtg cgatagcgcc cctgtgtgtt ctcgttatgt
     4081 tgaggaaaaa aataatggtt gctaagagat tcgaactctt gcatcttacg atacctgagt
     4141 attcccacag ttgggggatc tcgactctag ctagaggatc aattcgtaat catgtcatag
     4201 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     4261 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     4321 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4381 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4441 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4501 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4561 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4621 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4681 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     4741 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4801 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4861 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4921 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4981 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     5041 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     5101 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     5161 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     5221 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     5281 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5341 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5401 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5461 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5521 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5581 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5641 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     5701 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     5761 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5821 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5881 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5941 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     6001 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     6061 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     6121 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     6181 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     6241 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6301 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     6361 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     6421 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     6481 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     6541 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     6601 aacgcattta agcataaaca cgcactatgc cgttcttctc atgtatatat atatacaggc
     6661 aacacgcaga tataggtgcg acgtgaacag tgagctgtat gtgcgcagct cgcgttgcat
     6721 tttcggaagc gctcgttttc ggaaacgctt tgaagttcct attccgaagt tcctattctc
     6781 tagctagaaa gtataggaac ttcagagcgc ttttgaaaac caaaagcgct ctgaagacgc
     6841 actttcaaaa aaccaaaaac gcaccggact gtaacgagct actaaaatat tgcgaatacc
     6901 gcttccacaa acattgctca aaagtatctc tttgctatat atctctgtgc tatatcccta
     6961 tataacctac ccatccacct ttcgctcctt gaacttgcat ctaaactcga cctctacatt
     7021 ttttatgttt atctctagta ttactcttta gacaaaaaaa ttgtagtaag aactattcat
     7081 agagtgaatc gaaaacaata cgaaaatgta aacatttcct atacgtagta tatagagaca
     7141 aaatagaaga aaccgttcat aattttctga ccaatgaaga atcatcaacg ctatcacttt
     7201 ctgttcacaa agtatgcgca atccacatcg gtatagaata taatcgggga tgcctttatc
     7261 ttgaaaaaat gcacccgcag cttcgctagt aatcagtaaa cgcgggaagt ggagtcaggc
     7321 tttttttatg gaagagaaaa tagacaccaa agtagccttc ttctaacctt aacggaccta
     7381 cagtgcaaaa agttatcaag agactgcatt atagagcgca caaaggagaa aaaaagtaat
     7441 ctaagatgct ttgttagaaa aatagcgctc tcgggatgca tttttgtaga acaaaaaaga
     7501 agtatagatt ctttgttggt aaaatagcgc tctcgcgttg catttctgtt ctgtaaaaat
     7561 gcagctcaga ttctttgttt gaaaaattag cgctctcgcg ttgcattttt gttttacaaa
     7621 aatgaagcac agattcttcg ttggtaaaat agcgctttcg cgttgcattt ctgttctgta
     7681 aaaatgcagc tcagattctt tgtttgaaaa attagcgctc tcgcgttgca tttttgttct
     7741 acaaaatgaa gcacagatgc ttcgttgctt gcatgcaact tcttttcttt tt
//
