LOCUS       sequence-204897-        7342 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Prc1 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 7342)
  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 7342)
  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: pKP136 (pRS406-PRC1p-PRC1-PA-ADH1t)
FEATURES             Location/Qualifiers
     source          1..7342
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1827..2000
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             2001..2174
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAVIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     terminator      2210..2397
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2265..2289)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(2420..2436)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2421..2437)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2461..2480)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2462..2480)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2490..2507)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2490..2506)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2499..2521)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2647..3102
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2734..2753)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2944..2965
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3233..4036)
                     /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(4037..4252)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4276..4295)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4395..4417
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4455..4473)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4541..4645
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4646..4714,4715..5506)
                     /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(4864..4883)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5677..6265
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6166..6185
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6419..6436
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6553..6574
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6589..6594,6595..6612,6613..6619)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6627..6643
                     /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     6632..6654
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6651..6667
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6651..6667
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6686..6706
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6688..6706
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 acccgcgggg tctcaaagaa ggggcccact aataaaagca cgagataaga atgccagcaa
       61 aaaagctccg aaataattct tttcgtcttc cctcctagtc ttaacaagac aagagagaga
      121 gagagaaaga atactcacta gagattgttt cttttctact caacttaaag tatacatacg
      181 ctatgaaagc attcaccagt ttactatgtg gactaggcct gtccactaca ctcgctaagg
      241 ccatctcatt gcaaagaccg ttgggtctag ataaggacgt tttgctgcaa gctgcggaaa
      301 aatttggttt ggacctcgac ctggatcatc tcttgaagga gttggactcc aatgtattgg
      361 acgcttgggc ccaaatagag catttgtacc caaaccaggt tatgagcctt gaaacttcca
      421 ctaagccaaa attccctgaa gcaatcaaaa cgaagaaaga ctgggacttt gtggtcaaga
      481 atgacgcaat tgaaaactat cagcttcgtg tcaacaagat taaggaccct aaaatcctgg
      541 gcattgaccc aaatgtcaca cagtacacgg gttacttgga tgtggaagac gaggacaagc
      601 atttcttctt ttggactttt gaaagtagaa acgatcctgc aaaggatccg gtcatccttt
      661 ggttgaacgg gggtccaggt tgttcttcac taaccgggct gttctttgaa ttaggaccct
      721 catccattgg acctgatttg aaacccatcg ggaaccctta ctcttggaac agcaatgcca
      781 ccgtgatctt ccttgaccag cctgtcaacg ttgggttctc gtattccggg tcctcaggtg
      841 tttccaacac tgtcgccgct ggtaaggatg tctataactt cttggagttg ttcttcgatc
      901 agttccctga atacgtcaac aagggccaag atttccacat cgctggggaa tcctacgccg
      961 gccattacat ccctgttttt gcctctgaaa ttttgtctca caaggacaga aacttcaact
     1021 taacctccgt cttgatcgga aatggcctca ctgacccatt gactcagtat aactattacg
     1081 aaccaatggc ctgtggtgaa ggtggcgaac catctgtttt gccctcggag gaatgctctg
     1141 ctatggaaga ctctttggaa cgttgtttgg gcttgatcga gtcgtgctat gactcgcaat
     1201 ccgtctggtc ctgtgttcca gctaccattt attgtaataa cgcccaattg gctccttacc
     1261 aacgtaccgg cagaaacgtt tacgatatca ggaaggattg tgaaggtggc aatttgtgct
     1321 acccaacgtt acaagatatc gacgactact taaaccagga ctacgtcaaa gaagctgtcg
     1381 gtgcggaggt tgaccactac gaatcctgta acttcgatat caacagaaat ttcctgtttg
     1441 cgggtgattg gatgaagcct taccacaccg ccgtaacaga tcttttgaat caagacctac
     1501 ccattctggt atatgcaggc gataaagatt tcatctgtaa ctggttgggt aataaggcgt
     1561 ggacggatgt cttgccatgg aagtacgacg aagaatttgc aagccaaaaa gtacgtaact
     1621 ggactgcttc tatcaccgac gaggtcgctg gtgaagtcaa atcctacaag cacttcacct
     1681 atttgagagt cttcaatggt ggccacatgg ttccatttga cgtccctgaa aacgccttaa
     1741 gtatggttaa cgaatggatc cacggtggtt tctccttatc tagacggatc cccgggttaa
     1801 ttaagcttgc gcaacacgat gaagccgtgg acaacaaatt caacaaagaa caacaaaacg
     1861 cgttctatga gatcttacat ttacctaact taaacgaaga acaacgaaac gccttcatcc
     1921 aaagcttaaa agatgaccca agccaaagcg ctaacctttt agcagaagct aaaaagctaa
     1981 atgatgctca ggcgccgaaa gtagacaaca aattcaacaa agaacaacaa aacgcgttct
     2041 atgagatctt acatttacct aacttaaacg aagaacaacg aaacgccgtc atccaaagtt
     2101 taaaagatga cccaagccaa agcgctaacc ttttagcaga agctaaaaag ctaaatgatg
     2161 ctcaggcgcc gaaagtagac gcgaattaag gcgcgccact tctaaataag cgaatttctt
     2221 atgatttatg atttttatta ttaaataagt tataaaaaaa ataagtgtat acaaatttta
     2281 aagtgactct taggttttaa aacgaaaatt cttattcttg agtaactctt tcctgtaggt
     2341 caggttgctt tctcaggtat agtatgaggt cgctcttatt gaccacacct ctaccggcag
     2401 aattcgatat caagcttatc gataccgtcg acctcgaggg ggggcccggt acccaattcg
     2461 ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg
     2521 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     2581 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     2641 gaatggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     2701 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     2761 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     2821 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     2881 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     2941 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     3001 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     3061 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct
     3121 ccttacgcat ctgtgcggta tttcacaccg catagggtaa taactgatat aattaaattg
     3181 aagctctaat ttgtgagttt agtatacatg catttactta taatacagtt ttttagtttt
     3241 gctggccgca tcttctcaaa tatgcttccc agcctgcttt tctgtaacgt tcaccctcta
     3301 ccttagcatc ccttcccttt gcaaatagtc ctcttccaac aataataatg tcagatcctg
     3361 tagagaccac atcatccacg gttctatact gttgacccaa tgcgtctccc ttgtcatcta
     3421 aacccacacc gggtgtcata atcaaccaat cgtaaccttc atctcttcca cccatgtctc
     3481 tttgagcaat aaagccgata acaaaatctt tgtcgctctt cgcaatgtca acagtaccct
     3541 tagtatattc tccagtagat agggagccct tgcatgacaa ttctgctaac atcaaaaggc
     3601 ctctaggttc ctttgttact tcttctgccg cctgcttcaa accgctaaca atacctgggc
     3661 ccaccacacc gtgtgcattc gtaatgtctg cccattctgc tattctgtat acacccgcag
     3721 agtactgcaa tttgactgta ttaccaatgt cagcaaattt tctgtcttcg aagagtaaaa
     3781 aattgtactt ggcggataat gcctttagcg gcttaactgt gccctccatg gaaaaatcag
     3841 tcaagatatc cacatgtgtt tttagtaaac aaattttggg acctaatgct tcaactaact
     3901 ccagtaattc cttggtggta cgaacatcca atgaagcaca caagtttgtt tgcttttcgt
     3961 gcatgatatt aaatagcttg gcagcaacag gactaggatg agtagcagca cgttccttat
     4021 atgtagcttt cgacatgatt tatcttcgtt tcctgcaggt ttttgttctg tgcagttggg
     4081 ttaagaatac tgggcaattt catgtttctt caacactaca tatgcgtata tataccaatc
     4141 taagtctgtg ctccttcctt cgttcttcct tctgttcgga gattaccgaa tcaaaaaaat
     4201 ttcaaggaaa ccgaaatcaa aaaaaagaat aaaaaaaaaa tgatgaattg aaaaggtggt
     4261 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     4321 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     4381 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     4441 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     4501 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     4561 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     4621 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     4681 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     4741 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     4801 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     4861 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     4921 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     4981 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     5041 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     5101 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     5161 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     5221 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     5281 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     5341 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     5401 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     5461 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     5521 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     5581 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     5641 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     5701 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     5761 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     5821 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     5881 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     5941 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     6001 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     6061 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     6121 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     6181 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     6241 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     6301 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     6361 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     6421 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
     6481 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
     6541 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
     6601 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
     6661 ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac aaaagctgga
     6721 gctcgaatga gtgaaaaatt tgttagcaga gaagcaagaa ggaaagaagg aaaaaaaaaa
     6781 gaaaccgtgt aagaaatcat ggcgcctcaa aaaaaaaaaa atacgcgaaa tcgatttccg
     6841 tatatgatga tacatatgtt aggtctctta tggtagtttt tagggtctgt cctgtttttt
     6901 gaaggcattg gttagggtct agtaagtaac tcgtataaag agatgtactt ttatacggta
     6961 tgtgtggcgg ttatttccag tgtttttttt cccttatttt gtgggttcat ggaaatagta
     7021 catattgacc ttctgcacaa gaagccatat tgacagagca gtatgtgagg acctttcttc
     7081 tacacaatgt aacagtcaat ggttgtagtc aataccacga cctttacgtg cagttttaga
     7141 gcgaaacttc ggttttttga gacttacctc gtgtttgtgt ctccctggga atggagccgc
     7201 gccacccttg tctattatgt atagtcttat ttaatcatag cgatgttggt catccagtac
     7261 acttcggtag caaccttcgt ttgtgattgt cttggtaatt gcttccaaca actttatcca
     7321 tcattgagac aggggccata tc
//
