LOCUS       sequence-237454-        7255 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 7255)
  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 7255)
  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: pGBDU-Atg17(F317D)
FEATURES             Location/Qualifiers
     source          1..7255
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..404
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             432..872
                     /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"
     RBS             1943..1954
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      2554..2741
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2609..2633)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        2753..2964
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             2965..3768
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     primer_bind     complement(3912..3929)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4083..4671)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4163..4182)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4842..5633,5634..5702))
                     /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     5465..5484
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5703..5807)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5875..5893
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5931..5953)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6053..6072
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      6089..7253
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 ttgcatgcaa cttcttttct ttttttttct tttctctctc ccccgttgtt gtctcaccat
       61 atccgcaatg acaaaaaaat gatggaagac actaaaggaa aaaattaacg acaaagacag
      121 caccaacaga tgtcgttgtt ccagagctga tgaggggtat ctcgaagcac acgaaacttt
      181 ttccttcctt cattcacgca cactactctc taatgagcaa cggtatacgg ccttccttcc
      241 agttacttga atttgaaata aaaaaaagtt tgctgtcttg ctatcaagta taaatagacc
      301 tgcaattatt aatcttttgt ttcctcgtca ttgttctcgt tccctttctt ccttgtttct
      361 ttttctgcac aatatttcaa gctataccaa gcatacaatc aactccaagc ttgaagcaag
      421 cctcctgaaa gatgaagcta ctgtcttcta tcgaacaagc atgcgatatt tgccgactta
      481 aaaagctcaa gtgctccaaa gaaaaaccga agtgcgccaa gtgtctgaag aacaactggg
      541 agtgtcgcta ctctcccaaa accaaaaggt ctccgctgac tagggcacat ctgacagaag
      601 tggaatcaag gctagaaaga ctggaacagc tatttctact gatttttcct cgagaagacc
      661 ttgacatgat tttgaaaatg gattctttac aggatataaa agcattgtta acaggattat
      721 ttgtacaaga taatgtgaat aaagatgccg tcacagatag attggcttca gtggagactg
      781 atatgcctct aacattgaga cagcatagaa taagtgcgac atcatcatcg gaagagagta
      841 gtaacaaagg tcaaagacag ttgactgtat cgccggaatt ccccggggga tccatcgata
      901 acgaagcaga tgttacaaaa tttgttaata atgccaggaa aacgctgacc gatgctcaac
      961 ttttatgttc aagtgctaat ttaaggattg tagatattaa gaaaaaattg tcatcttggc
     1021 agttgagtat ttcaaaactc aattttctaa tagttggctt gagacagcaa ggaaagtttc
     1081 tttacactat tttaaaggaa ggcattggga caaagctaat ccagaaacaa tggaatcaag
     1141 ctgtcttagt ggtcttagtt gatgagatga agtactggca atatgaaatc acttctaaag
     1201 tacaaaggtt ggatggtata gtaaatgaat tgagcatatc agaaaaagat gataccgatc
     1261 cctctaaact aggagattac atctcgagag acaacgttaa cttattgaat gacaaactga
     1321 aagaagtgcc agtaattgag cgtcaaattg aaaacattaa gcttcaatac gaaaatatgg
     1381 tcagaaaagt taacaaagaa ctgattgata ccaagttgac ggacgtaact caaaaattcc
     1441 aaagtaaatt tggtatagat aacctgatgg aaacaaatgt tgcagagcag tttagcaggg
     1501 aactaacgga ccttgaaaaa gatttagcag agataatgaa ttcattgaca cagcactttg
     1561 ataagacatt gctgttacaa gataaaaaaa ttgataacga tgaacgtgag gagctgttta
     1621 aggtggtaca aggcgacgac aaagaactat acaacatttt caaaactctg catgaggtaa
     1681 ttgatgacgt ggacaaaaca attcttaact tgggtcaatt tttgcaggca aaaataaagg
     1741 aaaagacaga actacacagc gaagtttctg aaataataaa cgatttcaat agaaatttgg
     1801 aatatctatt aatctttaaa gatatttcca atctgattga tagcgacaaa aattcctgta
     1861 cacaagatat tcaaacaact aaggaacttt gtgaatttta tgataatttt gaagaaagct
     1921 acggtaactt agttctagaa gcaaagagga gaaaggatgt ggcaaacaga atgaaaacta
     1981 tattgaaaga ttgtgaaaag cagttgcaga atttagatgc tcaggaccag gaagaacgtc
     2041 agaattttat agcggaaaat ggaacttatc ttcctgagac aatctggccc ggtaaaattg
     2101 acgatttttc ttccctgtac actttaaatt acaacgtgaa gaatccttag aaaaaaggaa
     2161 aaaaaggctg cagagatcta tgaatcgtag atactgaaaa accccgcaag ttcacttcaa
     2221 ctgtgcatcg tgcaccatct caatttcttt catttataca tcgttttgcc ttcttttatg
     2281 taactatact cctctaagtt tcaatcttgg ccatgtaacc tctgatctat agaatttttt
     2341 aaatgactag aattaatgcc catctttttt ttggacctaa attcttcatg aaaatatatt
     2401 acgagggctt attcagaagc tttggacttc ttcgccagag gtttggtcaa gtctccaatc
     2461 aaggttgtcg gcttgtctac cttgccagaa atttacgaaa agatggaaaa gggtcaaatc
     2521 gttggtagat acgttgttga cacttctaaa taagcgaatt tcttatgatt tatgattttt
     2581 attattaaat aagttataaa aaaaataagt gtatacaaat tttaaagtga ctcttaggtt
     2641 ttaaaacgaa aattcttatt cttgagtaac tctttcctgt aggtcaggtt gctttctcag
     2701 gtatagcatg aggtcgctct tattgaccac acctctaccg gcatgccagc ttttcaattc
     2761 atcatttttt ttttattctt ttttttgatt tcggtttcct tgaaattttt ttgattcggt
     2821 aatctccgaa cagaaggaag aacgaaggaa ggagcacaga cttagattgg tatatatacg
     2881 catatgtagt gttgaagaaa catgaaattg cccagtattc ttaacccaac tgcacagaac
     2941 aaaaaccgga aacgaagata aatcatgtcg aaagctacat ataaggaacg tgctgctact
     3001 catcctagtc ctgttgctgc caagctattt aatatcatgc acgaaaagca aacaaacttg
     3061 tgtgcttcat tggatgttcg taccaccaag gaattactgg agttagttga agcattaggt
     3121 cccaaaattt gtttactaaa aacacatgtg gatatcttga ctgatttttc catggagggc
     3181 acagttaagc cgctaaaggc attatccgcc aagtacaatt ttttactctt cgaagacaga
     3241 aaatttgctg acattggtaa tacagtcaaa ttgcagtact ctgcgggtgt atacagaata
     3301 gcagaatggg cagacattac gaatgcacac ggtgtggtgg gcccaggtat tgttagcggt
     3361 ttgaagcagg cggcagaaga agtaacaaag gaacctagag gccttttgat gttagcagaa
     3421 ttgtcatgca agggctccct atctactgga gaatatacta agggtactgt tgacattgcg
     3481 aagagcgaca aagattttgt tatcggcttt attgctcaaa gagacatggg tggaagagat
     3541 gaaggttacg attggttgat tatgacaccc ggtgtgggtt tagatgacaa gggagacgca
     3601 ttgggtcaac agtatagaac cgtggatgat gtggtctcta caggatctga cattattatt
     3661 gttggaagag gactatttgc aaagggaagg gatgctaagg tagagggtga acgttacaga
     3721 aaagcaggct gggaagcata tttgagaaga tgcggccagc aaaactaaaa aactgtatta
     3781 taagtaaatg catgtatact aaactcacaa attagagctt caatttaatt atatcagtta
     3841 ttacccctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     3901 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     3961 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4021 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     4081 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4141 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4201 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4261 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4321 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4381 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4441 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4501 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     4561 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     4621 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     4681 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     4741 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     4801 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     4861 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     4921 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     4981 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5041 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5101 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5161 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5221 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5281 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5341 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5401 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5461 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     5521 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     5581 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     5641 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     5701 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     5761 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     5821 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     5881 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     5941 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     6001 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca
     6061 gagcagattg tactgagagt gcaccataac gcatttaagc ataaacacgc actatgccgt
     6121 tcttctcatg tatatatata tacaggcaac acgcagatat aggtgcgacg tgaacagtga
     6181 gctgtatgtg cgcagctcgc gttgcatttt cggaagcgct cgttttcgga aacgctttga
     6241 agttcctatt ccgaagttcc tattctctag ctagaaagta taggaacttc agagcgcttt
     6301 tgaaaaccaa aagcgctctg aagacgcact ttcaaaaaac caaaaacgca ccggactgta
     6361 acgagctact aaaatattgc gaataccgct tccacaaaca ttgctcaaaa gtatctcttt
     6421 gctatatatc tctgtgctat atccctatat aacctaccca tccacctttc gctccttgaa
     6481 cttgcatcta aactcgacct ctacattttt tatgtttatc tctagtatta ctctttagac
     6541 aaaaaaattg tagtaagaac tattcataga gtgaatcgaa aacaatacga aaatgtaaac
     6601 atttcctata cgtagtatat agagacaaaa tagaagaaac cgttcataat tttctgacca
     6661 atgaagaatc atcaacgcta tcactttctg ttcacaaagt atgcgcaatc cacatcggta
     6721 tagaatataa tcggggatgc ctttatcttg aaaaaatgca cccgcagctt cgctagtaat
     6781 cagtaaacgc gggaagtgga gtcaggcttt ttttatggaa gagaaaatag acaccaaagt
     6841 agccttcttc taaccttaac ggacctacag tgcaaaaagt tatcaagaga ctgcattata
     6901 gagcgcacaa aggagaaaaa aagtaatcta agatgctttg ttagaaaaat agcgctctcg
     6961 ggatgcattt ttgtagaaca aaaaagaagt atagattctt tgttggtaaa atagcgctct
     7021 cgcgttgcat ttctgttctg taaaaatgca gctcagattc tttgtttgaa aaattagcgc
     7081 tctcgcgttg catttttgtt ttacaaaaat gaagcacaga ttcttcgttg gtaaaatagc
     7141 gctttcgcgt tgcatttctg ttctgtaaaa atgcagctca gattctttgt ttgaaaaatt
     7201 agcgctctcg cgttgcattt ttgttctaca aaatgaagca cagatgcttc gttgc
//
