LOCUS       sequence-191859-        5819 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast integrative vector for expression of His-tagged POL30 (PCNA) 
            mutant K127/164R; Leu2 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5819)
  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 5819)
  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: YIp128-P30-His-pol30(K127/164R)
FEATURES             Location/Qualifiers
     source          1..5819
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             576..593
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1404..1591
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1459..1483)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(1615..1631)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1615..1631)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1628..1650)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1639..1655
                     /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(1663..1669,1670..1687,1688..1693))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1708..1729
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1846..1863)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2017..2605)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2097..2116)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2776..3567,3568..3636))
                     /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     3399..3418
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3637..3741)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3809..3827
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3829..4234
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             4235..5329
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     primer_bind     complement(5470..5492)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5592..5611
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(5805..5819,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(5819,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgaa ttcgccctta ttccttaact gctgaagaaa agcaacccta
       61 tgaatctaag gctcaagcag acaagaagag atacgaatct gaaaaggaat tgtacaatgc
      121 tacacgtgct tgatctaaca actcaatgtt ctctacctcc catacaacgt acttttgaaa
      181 cttcagaaag aagttgagta ggttgcatta tgtgttatta ttattactat tatcattatt
      241 ttacttatat atccttcttt tcttttatgt attctcttcc tttttcgcac aacttatgct
      301 gattcttcaa tatctaatta tttagcattt ttcttctcca tccgacgcac caaaacgcga
      361 aacgcgtaac tttttttttg gatttcaact gatagttttc gtactttgct tcctctggta
      421 cataaaatta tatataagaa acacttttgc tttagccttc ctttctttcc acttgcacct
      481 ttcactttcg ccgtcctttt tcactcacag caacaagcag caagcactaa gtacgcagtc
      541 aaaagagaga aaaaggatcc aatatgagag gatctcacca tcaccatcac catggcccgg
      601 gaaaaatgtt agaagcaaaa tttgaagaag catccctttt caagagaata attgatggtt
      661 tcaaagattg tgtccagttg gtcaatttcc aatgtaaaga agatggtatc attgcacaag
      721 ctgtcgatga ctcaagagtt ctattggtct ccttggaaat aggtgtcgaa gccttccaag
      781 aatatagatg tgaccatcct gttacgttag gtatggatct aacctcacta agtaaaatcc
      841 tacgttgtgg taacaacacc gatacattaa cactaattgc tgacaacaca ccggattcca
      901 tcatcttatt atttgaggat accaagaaag accgtatagc cgaatactct ctgaaattga
      961 tggatatcga tgctgatttc ttaagaattg aagaattaca gtacgactcc accctgtcat
     1021 tgccatcttc cgaattctct aaaattgttc gtgacttgtc ccaattgagt gattctatta
     1081 atatcatgat aaccagagaa acaataaagt ttgtagctga cggtgatatc ggatcaggtt
     1141 cagtcataat aaaaccattc gtggatatgg aacatcctga aacaagcatc aaacttgaaa
     1201 tggatcaacc tgtcgacttg acgttcggag ctaaatattt attggacatc attaagggct
     1261 cctccctttc tgatagagtt ggtatcaggc tctccagcga agctcctgct ttattccaat
     1321 ttgatttgaa gagtgggttc ctacagtttt tcttggctcc taaatttaat gacgaagaat
     1381 aactgcagcc aagctaattc cgggcgaatt tcttatgatt tatgattttt attattaaat
     1441 aagttataaa aaaaataagt gtatacaaat tttaaagtga ctcttaggtt ttaaaacgaa
     1501 aattcttatt cttgagtaac tctttcctgt aggtcaggtt gctttctcag gtatagcatg
     1561 aggtcgctct tattgaccac acctctaccg gcatgcaagc ttggcgtaat catggtcata
     1621 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     1681 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     1741 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     1801 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     1861 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     1921 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     1981 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     2041 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     2101 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     2161 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     2221 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     2281 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     2341 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     2401 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     2461 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     2521 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     2581 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     2641 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     2701 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     2761 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     2821 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     2881 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     2941 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     3001 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     3061 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     3121 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     3181 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     3241 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     3301 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     3361 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     3421 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     3481 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     3541 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     3601 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     3661 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     3721 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     3781 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccaa ctgtgggaat
     3841 actcaggtat cgtaagatgc aagagttcga atctcttagc aaccattatt tttttcctca
     3901 acataacgag aacacacagg ggcgctatcg cacagaatca aattcgatga ctggaaattt
     3961 tttgttaatt tcagaggtcg cctgacgcat ataccttttt caactgaaaa attgggagaa
     4021 aaaggaaagg tgagagcgcc ggaaccggct tttcatatag aatagagaag cgttcatgac
     4081 taaatgcttg catcacaata cttgaagttg acaatattat ttaaggacct attgtttttt
     4141 ccaataggtg gttagcaatc gtcttacttt ctaacttttc ttacctttta catttcagca
     4201 atatatatat atatttcaag gatataccat tctaatgtct gcccctaaga agatcgtcgt
     4261 tttgccaggt gaccacgttg gtcaagaaat cacagccgaa gccattaagg ttcttaaagc
     4321 tatttctgat gttcgttcca atgtcaagtt cgatttcgaa aatcatttaa ttggtggtgc
     4381 tgctatcgat gctacaggtg ttccacttcc agatgaggcg ctggaagcct ccaagaaggc
     4441 tgatgccgtt ttgttaggtg ctgtgggtgg tcctaaatgg ggtacaggta gtgttagacc
     4501 tgaacaaggt ttactaaaaa tccgtaaaga acttcaattg tacgccaact taagaccatg
     4561 taactttgca tccgactctc ttttagactt atctccaatc aagccacaat ttgctaaagg
     4621 tactgacttc gttgttgtca gagaattagt gggaggtatt tactttggta agagaaagga
     4681 agacgatggt gatggtgtcg cttgggatag tgaacaatac accgttccag aagtgcaaag
     4741 aatcacaaga atggccgctt tcatggccct acaacatgag ccaccattgc ctatttggtc
     4801 cttggataaa gctaatgttt tggcctcttc aagattatgg agaaaaactg tggaggaaac
     4861 catcaagaac gaatttccta cattgaaggt tcaacatcaa ttgattgatt ctgccgccat
     4921 gatcctagtt aagaacccaa cccacctaaa tggtattata atcaccagca acatgtttgg
     4981 tgatatcatc tccgatgaag cctccgttat cccaggttcc ttgggtttgt tgccatctgc
     5041 gtccttggcc tctttgccag acaagaacac cgcatttggt ttgtacgaac catgccacgg
     5101 ttctgctcca gatttgccaa agaataaggt caaccctatc gccactatct tgtctgctgc
     5161 aatgatgttg aaattgtcat tgaacttgcc tgaagaaggt aaggccattg aagatgcagt
     5221 taaaaaggtt ttggatgcag gtatcagaac tggtgattta ggtggttcca acagtaccac
     5281 cgaagtcggt gatgctgtcg ccgaagaagt taagaaaatc cttgcttaaa aagattctct
     5341 ttttttatga tatttgtaca taaactttat aaatgaaatt cataatagaa acgacacgaa
     5401 attacaaaat ggaatatgtt catagggtag gccctttcgt ctcgcgcgtt tcggtgatga
     5461 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
     5521 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg
     5581 gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat
     5641 accgcacaga tgcgtaagga gaaaataccg catcaggcgc cattcgccat tcaggctgcg
     5701 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
     5761 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgtt
//
