LOCUS       Exported                6253 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Yeast integrative vector for expression of His-tagged POL30 (PCNA) 
            mutant K127R; URA3 marker.
ACCESSION   .
VERSION     .
KEYWORDS    YIp211-P30-His-pol30(K127R)-3'
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6253)
  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 6253)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6253
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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(2548..2564)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2548..2564)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2561..2583)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2572..2588
                     /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(2596..2626)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2641..2662
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2779..2796)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2950..3538)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3030..3049)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3709..4569)
                     /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     4332..4351
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4570..4674)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4742..4760
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4767..4982
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             4983..5786
                     /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(5904..5926)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6026..6045
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6239..8
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6253..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 caccaaagaa 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 aaaaaaaaac
     1441 agtaaagttt gttttaaatg aaaataaata acaaagaaaa taaagactaa gatgcgttaa
     1501 atatcagctt tttgtgtgac ttatacagta tttatgacat atcttacatt aatctagaca
     1561 gcacataact tttgctatga acccttccaa atggaatatt gctttgtttc caagtgataa
     1621 tctttcgtca aaaacacttg agtattcaat tatggacgat ttattcgtgg tattcagggt
     1681 ttccacatcc aacaaagaaa acgttagttc ctttaagatg atattggcag gtatacaatg
     1741 agctagtaaa tcgtatagga cagctctgca ttcgattaaa gaattgacag atctctcttt
     1801 aacgattttc ctcgttaatt tatggatcac tataatccaa tctggtttta ttatagggct
     1861 actgcttttc aatgctaatt cgttgttcag tgccatagat tcaagcatta atagggagac
     1921 tcgcaagttt ccattcgatg cctgagcaat tctttttaaa atatcctttg tttctagttg
     1981 tattctttca tttgtcacca catcagacaa gatagttgaa atttcgctat cgcttggtgc
     2041 aggacaacga atcaacagac aacgggattt gataggagca attataggcg acatcgaatc
     2101 gcagaccatt atcaacctaa tgtttttgga gtatttttcc atggtacgtc ttaaagcagc
     2161 ttgagcatct tttgttaacg agttcgcctc gttgataata acacacttat atctatgggc
     2221 aagtccatcc ttagaatctt gaaagtccac ttgttccatt tgagccactt ctttcaatag
     2281 ttcttggatg acaattctat cattgttacc catatcactt ggcgtgatct ctaaatggta
     2341 tggcgagctg accacattca gttctagttt tctgttcgaa gcagtgacaa attgtctgac
     2401 atcaattttc aatctataga ctccaggtcc aaatatggac tccaataatg ccatacaacg
     2461 cgttttctta cctgtaccat ttggtccata cagtaaaaga tgaggtaaat cacgaggctg
     2521 caggcatgca agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     2581 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     2641 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     2701 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     2761 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     2821 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     2881 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     2941 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     3001 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     3061 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     3121 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     3181 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     3241 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     3301 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     3361 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     3421 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     3481 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     3541 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     3601 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     3661 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     3721 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     3781 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     3841 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     3901 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     3961 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     4021 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     4081 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     4141 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     4201 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     4261 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     4321 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     4381 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     4441 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     4501 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     4561 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     4621 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     4681 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     4741 aaataggcgt atcacgaggc cagcttttca attcatcatt ttttttttat tctttttttt
     4801 gatttcggtt tccttgaaat ttttttgatt cggtaatctc cgaacagaag gaagaacgaa
     4861 ggaaggagca cagacttaga ttggtatata tacgcatatg tagtgttgaa gaaacatgaa
     4921 attgcccagt attcttaacc caactgcaca gaacaaaaac atgcaggaaa cgaagataaa
     4981 tcatgtcgaa agctacatat aaggaacgtg ctgctactca tcctagtcct gttgctgcca
     5041 agctatttaa tatcatgcac gaaaagcaaa caaacttgtg tgcttcattg gatgttcgta
     5101 ccaccaagga attactggag ttagttgaag cattaggtcc caaaatttgt ttactaaaaa
     5161 cacatgtgga tatcttgact gatttttcca tggagggcac agttaagccg ctaaaggcat
     5221 tatccgccaa gtacaatttt ttactcttcg aagacagaaa atttgctgac attggtaata
     5281 cagtcaaatt gcagtactct gcgggtgtat acagaatagc agaatgggca gacattacga
     5341 atgcacacgg tgtggtgggc ccaggtattg ttagcggttt gaagcaggcg gcagaagaag
     5401 taacaaagga acctagaggc cttttgatgt tagcagaatt gtcatgcaag ggctccctat
     5461 ctactggaga atatactaag ggtactgttg acattgcgaa gagcgacaaa gattttgtta
     5521 tcggctttat tgctcaaaga gacatgggtg gaagagatga aggttacgat tggttgatta
     5581 tgacacccgg tgtgggttta gatgacaagg gagacgcatt gggtcaacag tatagaaccg
     5641 tggatgatgt ggtctctaca ggatctgaca ttattattgt tggaagagga ctatttgcaa
     5701 agggaaggga tgctaaggta gagggtgaac gttacagaaa agcaggctgg gaagcatatt
     5761 tgagaagatg cggccagcaa aactaaaaaa ctgtattata agtaaatgca tgtatactaa
     5821 actcacaaat tagagcttca atttaattat atcagttatt acccgccctt tcgtctcgcg
     5881 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     5941 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     6001 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     6061 atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag gcgccattcg
     6121 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
     6181 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc
     6241 cagtcacgac gtt
//
