LOCUS       sequence-192012-        6254 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast integrative vector for expression of His-tagged POL30 (PCNA) 
            mutant K127/164R; URA3 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6254)
  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 6254)
  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: YIp211-P30-His-pol30(K127/164R)-3'
FEATURES             Location/Qualifiers
     source          1..6254
                     /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"
     primer_bind     complement(2549..2565)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2549..2565)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2562..2584)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2573..2589
                     /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(2597..2603,2604..2621,2622..2627))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2642..2663
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2780..2797)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2951..3539)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3031..3050)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3710..4501,4502..4570))
                     /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     4333..4352
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4571..4675)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4743..4761
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4768..4983
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             4984..5787
                     /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"
     primer_bind     complement(5905..5927)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6027..6046
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(6240..6254,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(6254,1..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 aactgcagat gtaaattatc tatatagttg tatactaaaa ataataaaca aaaaaaaaaa
     1441 cagtaaagtt tgttttaaat gaaaataaat aacaaagaaa ataaagacta agatgcgtta
     1501 aatatcagct ttttgtgtga cttatacagt atttatgaca tatcttacat taatctagac
     1561 agcacataac ttttgctatg aacccttcca aatggaatat tgctttgttt ccaagtgata
     1621 atctttcgtc aaaaacactt gagtattcaa ttatggacga tttattcgtg gtattcaggg
     1681 tttccacatc caacaaagaa aacgttagtt cctttaagat gatattggca ggtatacaat
     1741 gagctagtaa atcgtatagg acagctctgc attcgattaa agaattgaca gatctctctt
     1801 taacgatttt cctcgttaat ttatggatca ctataatcca atctggtttt attatagggc
     1861 tactgctttt caatgctaat tcgttgttca gtgccataga ttcaagcatt aatagggaga
     1921 ctcgcaagtt tccattcgat gcctgagcaa ttctttttaa aatatccttt gtttctagtt
     1981 gtattctttc atttgtcacc acatcagaca agatagttga aatttcgcta tcgcttggtg
     2041 caggacaacg aatcaacaga caacgggatt tgataggagc aattataggc gacatcgaat
     2101 cgcagaccat tatcaaccta atgtttttgg agtatttttc catggtacgt cttaaagcag
     2161 cttgagcatc ttttgttaac gagttcgcct cgttgataat aacacactta tatctatggg
     2221 caagtccatc cttagaatct tgaaagtcca cttgttccat ttgagccact tctttcaata
     2281 gttcttggat gacaattcta tcattgttac ccatatcact tggcgtgatc tctaaatggt
     2341 atggcgagct gaccacattc agttctagtt ttctgttcga agcagtgaca aattgtctga
     2401 catcaatttt caatctatag actccaggtc caaatatgga ctccaataat gccatacaac
     2461 gcgttttctt acctgtacca tttggtccat acagtaaaag atgaggtaaa tcacgaggct
     2521 gcaggcatgc aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     2581 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     2641 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     2701 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     2761 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     2821 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     2881 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     2941 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3001 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3061 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3121 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     3181 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     3241 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     3301 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     3361 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     3421 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     3481 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     3541 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     3601 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     3661 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     3721 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     3781 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     3841 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     3901 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     3961 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4021 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4081 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4141 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     4201 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     4261 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     4321 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     4381 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     4441 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     4501 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     4561 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     4621 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     4681 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     4741 aaaataggcg tatcacgagg ccagcttttc aattcatcat ttttttttta ttcttttttt
     4801 tgatttcggt ttccttgaaa tttttttgat tcggtaatct ccgaacagaa ggaagaacga
     4861 aggaaggagc acagacttag attggtatat atacgcatat gtagtgttga agaaacatga
     4921 aattgcccag tattcttaac ccaactgcac agaacaaaaa catgcaggaa acgaagataa
     4981 atcatgtcga aagctacata taaggaacgt gctgctactc atcctagtcc tgttgctgcc
     5041 aagctattta atatcatgca cgaaaagcaa acaaacttgt gtgcttcatt ggatgttcgt
     5101 accaccaagg aattactgga gttagttgaa gcattaggtc ccaaaatttg tttactaaaa
     5161 acacatgtgg atatcttgac tgatttttcc atggagggca cagttaagcc gctaaaggca
     5221 ttatccgcca agtacaattt tttactcttc gaagacagaa aatttgctga cattggtaat
     5281 acagtcaaat tgcagtactc tgcgggtgta tacagaatag cagaatgggc agacattacg
     5341 aatgcacacg gtgtggtggg cccaggtatt gttagcggtt tgaagcaggc ggcagaagaa
     5401 gtaacaaagg aacctagagg ccttttgatg ttagcagaat tgtcatgcaa gggctcccta
     5461 tctactggag aatatactaa gggtactgtt gacattgcga agagcgacaa agattttgtt
     5521 atcggcttta ttgctcaaag agacatgggt ggaagagatg aaggttacga ttggttgatt
     5581 atgacacccg gtgtgggttt agatgacaag ggagacgcat tgggtcaaca gtatagaacc
     5641 gtggatgatg tggtctctac aggatctgac attattattg ttggaagagg actatttgca
     5701 aagggaaggg atgctaaggt agagggtgaa cgttacagaa aagcaggctg ggaagcatat
     5761 ttgagaagat gcggccagca aaactaaaaa actgtattat aagtaaatgc atgtatacta
     5821 aactcacaaa ttagagcttc aatttaatta tatcagttat tacccgccct ttcgtctcgc
     5881 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     5941 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     6001 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     6061 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
     6121 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     6181 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     6241 ccagtcacga cgtt
//
