LOCUS       sequence-191761-        5941 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast episomal vector for expression of His-tagged Smt3 (SUMO); TRP1
            marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5941)
  AUTHORS   Ulrich HD, Davies AA
  TITLE     In vivo detection and characterization of sumoylation targets in 
            Saccharomyces cerevisiae.
  JOURNAL   Methods Mol Biol. 2009;497:81-103. doi: 10.1007/978-1-59745-566-4_6.
  PUBMED    19107412
REFERENCE   2  (bases 1 to 5941)
  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"; journalName: "Methods Mol
            Biol."; date: "2009"; volume: "497:81-103. doi"; pages: " 
            10.1007/978-1-59745-566-4_6"
COMMENT     Sequence Label: YEp112-CUP1-His-Smt3
FEATURES             Location/Qualifiers
     source          1..5941
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..235
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
     CDS             282..302
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     CDS             309..602
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     terminator      636..823
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(691..715)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(847..863)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(847..863)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(860..882)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    871..887
                     /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(895..901,902..919,920..925))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    940..961
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1078..1095)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1249..1837)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1329..1348)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2008..2799,2800..2868))
                     /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     2631..2650
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2869..2973)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3041..3059
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3079..3180
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             3181..3855
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     rep_origin      3967..5313
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     primer_bind     5735..5757
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5749..5766
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5750..5766
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ctagttagaa aaagacattt ttgctgtcag tcactgtcaa gagattcttt tgctggcatt
       61 tcttctagaa gcaaaaagag cgatgcgtct tttccgctga accgttccag caaaaaagac
      121 taccaacgca atatggattg tcagaatcat ataaaagaga agcaaataac tccttgtctt
      181 gtatcaattg cattataata tcttcttgtt agtgcaatat catatagaag tcatcgaaat
      241 agatattaag aaaaacaaac tgtaacggat ccattatggg ccatcatcac catcatcacc
      301 atagcggcat gtcggactca gaagtcaatc aagaagctaa gccagaggtc aagccagaag
      361 tcaagcctga gactcacatc aatttaaagg tgtccgatgg atcttcagag atcttcttca
      421 agatcaaaaa gaccactcct ttaagaaggc tgatggaagc gttcgctaaa agacagggta
      481 aggaaatgga ctccttaaga ttcttgtacg acggtattag aattcaagct gatcagaccc
      541 ctgaagattt ggacatggag gataacgata ttattgaggc tcacagagaa cagattggtg
      601 gtgctacgta ttagctgcag ccaagctaat tccgggcgaa tttcttatga tttatgattt
      661 ttattattaa ataagttata aaaaaaataa gtgtatacaa attttaaagt gactcttagg
      721 ttttaaaacg aaaattctta ttcttgagta actctttcct gtaggtcagg ttgctttctc
      781 aggtatagca tgaggtcgct cttattgacc acacctctac cggcatgcaa gcttggcgta
      841 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
      901 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
      961 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     1021 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     1081 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     1141 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     1201 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     1261 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     1321 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     1381 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     1441 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     1501 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     1561 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     1621 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     1681 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     1741 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     1801 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     1861 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     1921 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     1981 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     2041 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     2101 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     2161 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     2221 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     2281 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     2341 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     2401 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     2461 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     2521 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     2581 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     2641 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     2701 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     2761 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     2821 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     2881 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     2941 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     3001 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     3061 ctttcgtctt caagaattaa ttcggtcgaa aaaagaaaag gagagggcca agagggaggg
     3121 cattggtgac tattgagcac gtgagtatac gtgattaagc acacaaaggc agcttggagt
     3181 atgtctgtta ttaatttcac aggtagttct ggtccattgg tgaaagtttg cggcttgcag
     3241 agcacagagg ccgcagaatg tgcactagat tccgatgctg acttgctggg tattatatgt
     3301 gtgcccaata gaaagagaac aattgacccg gttattgcaa ggaaaatttc aagtcttgta
     3361 aaagcatata aaaatagttc aggcactccg aaatacttgg ttggcgtgtt tcgtaatcaa
     3421 cctaaggagg atgttttggc tctggtcaat gattacggca ttgatatcgt ccaactgcat
     3481 ggagatgagt cgtggcaaga ataccaagag ttcctcggtt tgccagttat taaaagactc
     3541 gtatttccaa aagactgcaa catactactc agtgcagctt cacagaaacc tcattcgttt
     3601 attcccttgt ttgattcaga agcaggtggg acaggtgaac ttttggattg gaactcgatt
     3661 tctgactggg ttggaaggca agagagcccc gagagcttac attttatgtt agctggtgga
     3721 ctgacgccag aaaatgttgg tgatgcgctt agattaaatg gcgttattgg tgttgatgta
     3781 agcggaggtg tggagacaaa tggtgtaaaa gactctaaca aaatagcaaa tttcgtcaaa
     3841 aatgctaaga aataggttat tactgagtag tatttattta agtattgttt gtgcacttgc
     3901 ctgcaagcct tttgaaaagc aagcataaaa gatcaattct catgtttgac agcttatcat
     3961 cggatcgatc caatatcaaa ggaaatgata gcattgaagg atgagactaa tccaattgag
     4021 gagtggcagc atatagaaca gctaaagggt agtgctgaag gaagcatacg ataccccgca
     4081 tggaatggga taatatcaca ggaggtacta gactaccttt catcctacat aaatagacgc
     4141 atataagtac gcatttaagc ataaacacgc actatgccgt tcttctcatg tatatatata
     4201 tacaggcaac acgcagatat aggtgcgacg tgaacagtga gctgtatgtg cgcagctcgc
     4261 gttgcatttt cggaagcgct cgttttcgga aacgctttga agttcctatt ccgaagttcc
     4321 tattctctag ctagaaagta taggaacttc agagcgcttt tgaaaaccaa aagcgctctg
     4381 aagacgcact ttcaaaaaac caaaaacgca ccggactgta acgagctact aaaatattgc
     4441 gaataccgct tccacaaaca ttgctcaaaa gtatctcttt gctatatatc tctgtgctat
     4501 atccctatat aacctaccca tccacctttc gctccttgaa cttgcatcta aactcgacct
     4561 ctacattttt tatgtttatc tctagtatta ctctttagac aaaaaaattg tagtaagaac
     4621 tattcataga gtgaatcgaa aacaatacga aaatgtaaac atttcctata cgtagtatat
     4681 agagacaaaa tagaagaaac cgttcataat tttctgacca atgaagaatc atcaacgcta
     4741 tcactttctg ttcacaaagt atgcgcaatc cacatcggta tagaatataa tcggggatgc
     4801 ctttatcttg aaaaaatgca cccgcagctt cgctagtaat cagtaaacgc gggaagtgga
     4861 gtcaggcttt ttttatggaa gagaaaatag acaccaaagt agccttcttc taaccttaac
     4921 ggacctacag tgcaaaaagt tatcaagaga ctgcattata gagcgcacaa aggagaaaaa
     4981 aagtaatcta agatgctttg ttagaaaaat agcgctctcg ggatgcattt ttgtagaaca
     5041 aaaaagaagt atagattctt tgttggtaaa atagcgctct cgcgttgcat ttctgttctg
     5101 taaaaatgca gctcagattc tttgtttgaa aaattagcgc tctcgcgttg catttttgtt
     5161 ttacaaaaat gaagcacaga ttcttcgttg gtaaaatagc gctttcgcgt tgcatttctg
     5221 ttctgtaaaa atgcagctca gattctttgt ttgaaaaatt agcgctctcg cgttgcattt
     5281 ttgttctaca aaatgaagca cagatgcttc gttaacaaag atatgctatt gaagtgcaag
     5341 atggaaacgc agaaaatgaa ccggggatgc gacgtgcaag attacctatg caatagatgc
     5401 aatagtttct ccaggaaccg aaatacatac attgtcttcc gtaaagcgct agactatata
     5461 ttattataca ggttcaaata tactatctgt ttcagggaaa actcccaggt tcggatgttc
     5521 aaaattcaat gatgggtaac aagtacgatc cgatatatgc ggtgtgaaat accgcacaga
     5581 tgcgtaagga gaaaataccg catcaggcgc cattcgccat tcaggctgcg caactgttgg
     5641 gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct
     5701 gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg
     5761 gccagtgaat tcgagctccc attaccgaca tttgggcgct atacgtgcat atgttcatgt
     5821 atgtatctgt atttaaaaca cttttgtatt atttttcctc atatatgtgt ataggtttat
     5881 acggatgatt taattattac ttcaccaccc tttatttcag gctgatatct tagccttgtt
     5941 a
//
