LOCUS       sequence-204907-        7270 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Atg42/Ybr139w with a C-terminal PA tag in yeast cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7270)
  AUTHORS   Parzych KR, Ariosa A, Mari M, Klionsky DJ
  TITLE     A newly characterized vacuolar serine carboxypeptidase, 
            Atg42/Ybr139w, is required for normal vacuole function and the 
            terminal steps of autophagy in the yeast Saccharomyces cerevisiae.
  JOURNAL   Mol Biol Cell. 2018 May 1;29(9):1089-1099. doi: 
            10.1091/mbc.E17-08-0516. Epub 2018 Mar 22.
  PUBMED    29514932
REFERENCE   2  (bases 1 to 7270)
  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: 
            "10.1091/mbc.E17-08-0516"; journalName: "Mol Biol Cell"; date: 
            "2018-05-1- 1"; volume: "29"; issue: "9"; pages: "1089-1099"
COMMENT     Sequence Label: pKP135 (pRS406-YBR139Wp-YBR139W-PA-ADH1t)
FEATURES             Location/Qualifiers
     source          1..7270
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    132..153
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(168..173,174..191,192..198)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    206..222
                     /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)."
     primer_bind     211..233
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     230..246
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     230..246
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     265..285
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        267..285
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             2676..2849
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             2850..3023
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAVIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     terminator      3059..3246
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(3114..3138)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(3269..3285)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3270..3286)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(3310..3329)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3311..3329)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3339..3356)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3339..3355)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3348..3370)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3496..3951
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3583..3602)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3793..3814
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4082..4885)
                     /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"
     promoter        complement(4886..5101)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(5125..5144)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5244..5266
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5304..5322)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5390..5494
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5495..5563,5564..6355)
                     /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     complement(5713..5732)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6526..7114
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7015..7034
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     join(7268..7270,1..15)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
       61 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
      121 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
      181 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
      241 tatgaccatg attacgccaa gcgcgcaatt aaccctcact aaagggaaca aaagctggag
      301 ctcttgcatg gaggcgcaat tggcaaagac gtaaagctct tgtcatcaat tagatctaat
      361 ttggaaacag attggtacaa tttggatgga attttaggct tggcacctat caattgtgag
      421 cgacgctttc ttacattcaa taactgttgg ttctgatttt cactttgact attgccttgg
      481 tatggagaat ttctctgcaa tggttttgcc tgcttcgaag gagtgctgtt actatcgaag
      541 ataaattcct tcttttcaaa ttgccattgt aacatggagt tattctctat atcctggacg
      601 actggaggag tcaagggcct ggaaagtacg gatcctgtag aaatatcact ggcaattata
      661 ctgagtttat tgttggtgtt gctattcata tttgagctgg tattggacga aagcaaggat
      721 aatgagtttg tgtctactga ctccaacacc ctaggctgga tttgatcttt ctccatcctt
      781 taattttaac cttttaatta gtggttggat caagttttcg agactattcc aatctgtgac
      841 ttgttggata aatagttttt gactcgttta gtataatcct tttttctaaa agtgcttaga
      901 gttctctaag atgttcttgt ttacaatgtg agcgatttag gaaatttcct aaaaatggcc
      961 gaggcggcgc tagcattcta cgaaaggtga gataacgctt cgttatcgaa aaatgtcagg
     1021 ggacaggggt tatataagaa cgaaaattgt catcctgcat tttttcttta aaacagctat
     1081 acaaaaagtg ataccgacat acaatgaagt atctaaactt agttttcgtg cttcagcttc
     1141 ttattagcat caaatacgcc tcattcggcc gagccttttc tctttttgaa gatgatacca
     1201 cctttgccaa tttggataaa cagctaaagc ttccacagaa tacacagcaa acccttaaat
     1261 tggaccgttt gaatcacgat gatccgctgt ttacaacttt tatttcttct gtggacacag
     1321 attacagttt gagacttaga acagtagatc cttctaaact aggaattgac accgtaaaac
     1381 aatggtcggg ttacatggac tataaggatt ccaaacactt tttttactgg ttttttgaaa
     1441 gtaggaacga tcctgctaac gacccaatta ttctttggtt aaatggtgga cctggttgtt
     1501 cctcgtttac tgggttgcta tttgaactag gcccctcatc aattggcgcc gatatgaaac
     1561 caatccacaa tccctattct tggaataata acgcttcaat gatcttctta gaacagccac
     1621 tcggagtcgg cttttcctat ggtgatgaaa aagtctcctc tacaaaatta gcaggcaaag
     1681 atgcgtacat tttcctggaa ttgttttttg aagcttttcc tcatttacgc tccaacgatt
     1741 tccacattgc aggcgaatcc tatgcaggac attatatccc tcaaattgca catgagatcg
     1801 ttgtcaagaa ccctgaaaga acgttcaatt taacttcagt tatgattggt aatggtatca
     1861 cagacccttt gattcaagca gattattatg aaccaatggc atgcgggaaa gggggctatc
     1921 accctgttct ctcatcagaa gaatgtgaga aaatgagtaa agctgcaggt cgttgtcgta
     1981 ggttgaacaa gttatgttat gcttctaaat caagtttacc atgcatagtc gccactgctt
     2041 actgtgactc tgcacttttg gaaccgtaca ttaacacagg actcaacgtc tatgacatta
     2101 gagggccctg tgaagataat agtactgatg gtatgtgtta tacaggtctc cgctatgtcg
     2161 accagtatat gaattttcct gaagttcaag aaacgctagg gtccgacgtg cataattatt
     2221 ctggctgtga taatgacgtg ttcaccggat ttttgtttac gggcgatgga agtaaaccat
     2281 ttcaacaata tattgctgaa ttattaaatc acaacattcc ggtattaata tatgcgggtg
     2341 ataaggatta tatttgtaat tggctgggaa accatgcttg gtccaatgag ttggaatgga
     2401 tcaataaacg taggtatcag agaaggatgt taagaccatg ggtcagtaaa gaaacaggtg
     2461 aagagttggg acaagtcaag aactatggcc ctttcacctt tttgagaata tacgatgccg
     2521 gtcatatggt gccctatgat caaccggagg caagtttgga aatggtcaac agttggattt
     2581 ccggtaatcg tgctttttcg gatctttcca ccttggaaaa tgctagttct agacggatcc
     2641 ccgggttaat taagcttgcg caacacgatg aagccgtgga caacaaattc aacaaagaac
     2701 aacaaaacgc gttctatgag atcttacatt tacctaactt aaacgaagaa caacgaaacg
     2761 ccttcatcca aagcttaaaa gatgacccaa gccaaagcgc taacctttta gcagaagcta
     2821 aaaagctaaa tgatgctcag gcgccgaaag tagacaacaa attcaacaaa gaacaacaaa
     2881 acgcgttcta tgagatctta catttaccta acttaaacga agaacaacga aacgccgtca
     2941 tccaaagttt aaaagatgac ccaagccaaa gcgctaacct tttagcagaa gctaaaaagc
     3001 taaatgatgc tcaggcgccg aaagtagacg cgaattaagg cgcgccactt ctaaataagc
     3061 gaatttctta tgatttatga tttttattat taaataagtt ataaaaaaaa taagtgtata
     3121 caaattttaa agtgactctt aggttttaaa acgaaaattc ttattcttga gtaactcttt
     3181 cctgtaggtc aggttgcttt ctcaggtata gtatgaggtc gctcttattg accacacctc
     3241 taccggcaga attcgatatc aagcttatcg ataccgtcga cctcgagggg gggcccggta
     3301 cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt
     3361 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     3421 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     3481 ctgaatggcg aatggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     3541 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     3601 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     3661 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     3721 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     3781 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     3841 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     3901 tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt tcctgatgcg
     3961 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atagggtaat aactgatata
     4021 attaaattga agctctaatt tgtgagttta gtatacatgc atttacttat aatacagttt
     4081 tttagttttg ctggccgcat cttctcaaat atgcttccca gcctgctttt ctgtaacgtt
     4141 caccctctac cttagcatcc cttccctttg caaatagtcc tcttccaaca ataataatgt
     4201 cagatcctgt agagaccaca tcatccacgg ttctatactg ttgacccaat gcgtctccct
     4261 tgtcatctaa acccacaccg ggtgtcataa tcaaccaatc gtaaccttca tctcttccac
     4321 ccatgtctct ttgagcaata aagccgataa caaaatcttt gtcgctcttc gcaatgtcaa
     4381 cagtaccctt agtatattct ccagtagata gggagccctt gcatgacaat tctgctaaca
     4441 tcaaaaggcc tctaggttcc tttgttactt cttctgccgc ctgcttcaaa ccgctaacaa
     4501 tacctgggcc caccacaccg tgtgcattcg taatgtctgc ccattctgct attctgtata
     4561 cacccgcaga gtactgcaat ttgactgtat taccaatgtc agcaaatttt ctgtcttcga
     4621 agagtaaaaa attgtacttg gcggataatg cctttagcgg cttaactgtg ccctccatgg
     4681 aaaaatcagt caagatatcc acatgtgttt ttagtaaaca aattttggga cctaatgctt
     4741 caactaactc cagtaattcc ttggtggtac gaacatccaa tgaagcacac aagtttgttt
     4801 gcttttcgtg catgatatta aatagcttgg cagcaacagg actaggatga gtagcagcac
     4861 gttccttata tgtagctttc gacatgattt atcttcgttt cctgcaggtt tttgttctgt
     4921 gcagttgggt taagaatact gggcaatttc atgtttcttc aacactacat atgcgtatat
     4981 ataccaatct aagtctgtgc tccttccttc gttcttcctt ctgttcggag attaccgaat
     5041 caaaaaaatt tcaaggaaac cgaaatcaaa aaaaagaata aaaaaaaaat gatgaattga
     5101 aaaggtggta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     5161 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     5221 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     5281 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     5341 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     5401 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     5461 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     5521 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     5581 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     5641 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     5701 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     5761 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     5821 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     5881 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     5941 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     6001 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     6061 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     6121 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     6181 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     6241 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     6301 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     6361 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     6421 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     6481 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     6541 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     6601 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     6661 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     6721 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     6781 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     6841 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     6901 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     6961 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     7021 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     7081 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     7141 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     7201 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     7261 cgagcgcagc
//
