LOCUS       sequence-191768-        6714 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast episomal vector for expression of His-tagged ubiquitin; HIS3 
            marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6714)
  AUTHORS   Davies AA, Ulrich HD
  TITLE     Detection of PCNA modifications in Saccharomyces cerevisiae.
  JOURNAL   Methods Mol Biol. 2012;920:543-67. doi: 
            10.1007/978-1-61779-998-3_36.
  PUBMED    22941627
REFERENCE   2  (bases 1 to 6714)
  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: "2012"; volume: "920:543-67. doi"; pages: " 
            10.1007/978-1-61779-998-3_36"
COMMENT     Sequence Label: pRS423-CUP1-His-Ubi
FEATURES             Location/Qualifiers
     source          1..6714
                     /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..536
                     /codon_start=1
                     /product="Saccharomyces cerevisiae ubiquitin"
                     /label=ubiquitin
                     /translation="MQIFVKTLTGKTITLEVESSDTIDNVKSKIQDKEGIPPDQQRLIF
                     AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     terminator      561..748
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(616..640)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(763..779)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(763..779)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(816..836)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(816..834)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(855..871)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(855..871)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(868..890)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    879..895
                     /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(903..909,910..927,928..933))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    948..969
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1086..1103)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1257..1845)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1337..1356)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2016..2807,2808..2876))
                     /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     2639..2658
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2877..2981)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3008..4350
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(3971..4018)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     4391..4409
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4447..4469)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4569..4588
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4735..4922
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             4923..5585
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(5845..6300)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5982..6003)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6194..6213
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6424..6446
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6438..6455
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6439..6455
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6465..6484
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6465..6483
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     6508..6524
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     6509..6525
                     /label=KS primer
                     /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 gcaaattttt gtcaagactt taaccggtaa gactattacc ctggaagttg
      361 aatcttctga cactattgac aatgtcaagt ccaagatcca agacaaggaa ggtattccac
      421 ctgaccaaca aagattgatc tttgctggta agcaattgga agatggtaga actttgtccg
      481 actacaacat ccaaaaggaa tctactctac acttggtctt gagattgaga ggtggttgac
      541 tgcagccaag ctaattccgg gcgaatttct tatgatttat gatttttatt attaaataag
      601 ttataaaaaa aataagtgta tacaaatttt aaagtgactc ttaggtttta aaacgaaaat
      661 tcttattctt gagtaactct ttcctgtagg tcaggttgct ttctcaggta tagcatgagg
      721 tcgctcttat tgaccacacc tctaccggca tgcaagctgg gggatccact agttctagag
      781 cggccgccac cgcggtggag ctccagcttt tgttcccttt agtgagggtt aattgcgcgc
      841 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
      901 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
      961 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     1021 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     1081 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     1141 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     1201 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     1261 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     1321 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     1381 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     1441 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     1501 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     1561 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     1621 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     1681 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     1741 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     1801 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     1861 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     1921 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     1981 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     2041 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     2101 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     2161 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     2221 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     2281 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     2341 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     2401 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     2461 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     2521 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     2581 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     2641 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     2701 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     2761 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     2821 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     2881 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     2941 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     3001 ccacctgaac gaagcatctg tgcttcattt tgtagaacaa aaatgcaacg cgagagcgct
     3061 aatttttcaa acaaagaatc tgagctgcat ttttacagaa cagaaatgca acgcgaaagc
     3121 gctattttac caacgaagaa tctgtgcttc atttttgtaa aacaaaaatg caacgcgaga
     3181 gcgctaattt ttcaaacaaa gaatctgagc tgcattttta cagaacagaa atgcaacgcg
     3241 agagcgctat tttaccaaca aagaatctat acttcttttt tgttctacaa aaatgcatcc
     3301 cgagagcgct atttttctaa caaagcatct tagattactt tttttctcct ttgtgcgctc
     3361 tataatgcag tctcttgata actttttgca ctgtaggtcc gttaaggtta gaagaaggct
     3421 actttggtgt ctattttctc ttccataaaa aaagcctgac tccacttccc gcgtttactg
     3481 attactagcg aagctgcggg tgcatttttt caagataaag gcatccccga ttatattcta
     3541 taccgatgtg gattgcgcat actttgtgaa cagaaagtga tagcgttgat gattcttcat
     3601 tggtcagaaa attatgaacg gtttcttcta ttttgtctct atatactacg tataggaaat
     3661 gtttacattt tcgtattgtt ttcgattcac tctatgaata gttcttacta caattttttt
     3721 gtctaaagag taatactaga gataaacata aaaaatgtag aggtcgagtt tagatgcaag
     3781 ttcaaggagc gaaaggtgga tgggtaggtt atatagggat atagcacaga gatatatagc
     3841 aaagagatac ttttgagcaa tgtttgtgga agcggtattc gcaatatttt agtagctcgt
     3901 tacagtccgg tgcgtttttg gttttttgaa agtgcgtctt cagagcgctt ttggttttca
     3961 aaagcgctct gaagttccta tactttctag agaataggaa cttcggaata ggaacttcaa
     4021 agcgtttccg aaaacgagcg cttccgaaaa tgcaacgcga gctgcgcaca tacagctcac
     4081 tgttcacgtc gcacctatat ctgcgtgttg cctgtatata tatatacatg agaagaacgg
     4141 catagtgcgt gtttatgctt aaatgcgtac ttatatgcgt ctatttatgt aggatgaaag
     4201 gtagtctagt acctcctgtg atattatccc attccatgcg gggtatcgta tgcttccttc
     4261 agcactaccc tttagctgtt ctatatgctg ccactcctca attggattag tctcatcctt
     4321 caatgctatc atttcctttg atattggatc atactaagaa accattatta tcatgacatt
     4381 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     4441 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     4501 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     4561 taactatgcg gcatcagagc agattgtact gagagtgcac cataaattcc cgttttaaga
     4621 gcttggtgag cgctaggagt cactgccagg tatcgtttga acacggcatt agtcagggaa
     4681 gtcataacac agtcctttcc cgcaattttc tttttctatt actcttggcc tcctctagta
     4741 cactctatat ttttttatgc ctcggtaatg attttcattt ttttttttcc acctagcgga
     4801 tgactctttt tttttcttag cgattggcat tatcacataa tgaattatac attatataaa
     4861 gtaatgtgat ttcttcgaag aatatactaa aaaatgagca ggcaagataa acgaaggcaa
     4921 agatgacaga gcagaaagcc ctagtaaagc gtattacaaa tgaaaccaag attcagattg
     4981 cgatctcttt aaagggtggt cccctagcga tagagcactc gatcttccca gaaaaagagg
     5041 cagaagcagt agcagaacag gccacacaat cgcaagtgat taacgtccac acaggtatag
     5101 ggtttctgga ccatatgata catgctctgg ccaagcattc cggctggtcg ctaatcgttg
     5161 agtgcattgg tgacttacac atagacgacc atcacaccac tgaagactgc gggattgctc
     5221 tcggtcaagc ttttaaagag gccctagggg ccgtgcgtgg agtaaaaagg tttggatcag
     5281 gatttgcgcc tttggatgag gcactttcca gagcggtggt agatctttcg aacaggccgt
     5341 acgcagttgt cgaacttggt ttgcaaaggg agaaagtagg agatctctct tgcgagatga
     5401 tcccgcattt tcttgaaagc tttgcagagg ctagcagaat taccctccac gttgattgtc
     5461 tgcgaggcaa gaatgatcat caccgtagtg agagtgcgtt caaggctctt gcggttgcca
     5521 taagagaagc cacctcgccc aatggtacca acgatgttcc ctccaccaaa ggtgttctta
     5581 tgtagtgaca ccgattattt aaagctgcag catacgatat atatacatgt gtatatatgt
     5641 atacctatga atgtcagtaa gtatgtatac gaacagtatg atactgaaga tgacaaggta
     5701 atgcatcatt ctatacgtgt cattctgaac gaggcgcgct ttcctttttt ctttttgctt
     5761 tttctttttt tttctcttga actcgacgga tctatgcggt gtgaaatacc gcacagatgc
     5821 gtaaggagaa aataccgcat caggaaattg taagcgttaa tattttgtta aaattcgcgt
     5881 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     5941 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     6001 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     6061 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     6121 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     6181 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     6241 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     6301 ccattcgcca ttcaggctgc gcaactgttg ggaagggcgc ggtgcgggcc tcttcgctat
     6361 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
     6421 tttcccagtc acgacgttgt aaaacgacgg ccagtgagcg cgcgtaatac gactcactat
     6481 agggcgaatt gggtaccggg ccccccctcg aggtcgacgg tatcgataag cttgatatcg
     6541 aattcgagct cccattaccg acatttgggc gctatacgtg catatgttca tgtatgtatc
     6601 tgtatttaaa acacttttgt attatttttc ctcatatatg tgtataggtt tatacggatg
     6661 atttaattat tacttcacca ccctttattt caggctgata tcttagcctt gtta
//
