LOCUS       sequence-15-expo        6860 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6860)
  AUTHORS   Schirmer EC, Homann OR, Kowal AS, Lindquist S
  TITLE     Dominant gain-of-function mutations in Hsp104p reveal crucial roles 
            for the middle region.
  JOURNAL   Mol Biol Cell 2004 May;15(5):2061-72. Epub 2004 Feb 20.
  PUBMED    14978213
REFERENCE   2  (bases 1 to 6860)
  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: "Mol Biol 
            Cell"; date: "2004-05"; volume: "15(5)"; pages: "2061-72. Epub 2004 
            Feb 20"
COMMENT     Sequence Label: GAL-CDC12(K351N)
FEATURES             Location/Qualifiers
     source          1..6860
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    70..573
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        610..714
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(715..783,784..1575)
                     /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     complement(933..952)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1746..2334
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2235..2254
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2488..2505
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2658..2663,2664..2681,2682..2688)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2696..2712
                     /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)."
     primer_bind     2701..2723
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2720..2736
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2720..2736
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2757..2775
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     2804..2820
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2805..2821
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2846..3510
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     protein_bind    3062..3179
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3474..3497
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     complement(4868..4887)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4869..4887)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4894..4911)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4894..4910)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4903..4925)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5055..5510
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5142..5161)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5352..5373
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5641..6444)
                     /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"
     promoter        complement(6445..6665)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(6691..6710)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6810..6832
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttaggacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa tgatggaata
      121 atttgggaat ttactctgtg tttatttatt tttatgtttt gtatttggat tttagaaagt
      181 aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa
      241 atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc agattaaata
      301 gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg tgtgtggtct
      361 tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa gagcaagata
      421 aaaggtagta tttgttggcg atccccctag agtcttttac atcttcggaa aacaaaaact
      481 attttttctt taatttcttt ttttactttc tatttttaat ttatatattt atattaaaaa
      541 atttaaatta taattatttt tatagcacgt gatgaaaagg acccaggtgg cacttttcgg
      601 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
      661 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
      721 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
      781 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
      841 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
      901 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
      961 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     1021 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     1081 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     1141 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     1201 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     1261 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     1321 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     1381 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     1441 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     1501 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     1561 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     1621 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     1681 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     1741 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     1801 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     1861 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     1921 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     1981 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     2041 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     2101 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     2161 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     2221 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     2281 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     2341 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     2401 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     2461 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     2521 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     2581 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttacctcact
     2641 cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg
     2701 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gctcggaatt
     2761 aaccctcact aaagggaaca aaagctgggt accgggcccc ccctcgaggt cgacggtatc
     2821 gataagcttg atatcgaatt cgaattttca aaaattctta cttttttttt ggatggacgc
     2881 aaagaagttt aataatcata ttacatggca ttaccaccat atacatatcc atatacatat
     2941 ccatatctaa tcttacttat atgttgtgga aatgtaaaga gccccattat cttagcctaa
     3001 aaaaaccttc tctttggaac tttcagtaat acgcttaact gctcattgct atattgaagt
     3061 acggattaga agccgccgag cgggtgacag ccctccgaag gaagactctc ctccgtgcgt
     3121 cctcgtcttc accggtcgcg ttcctgaaac gcagatgtgc ctcgcgccgc actgctccga
     3181 acaataaaga ttctacaata ctagctttta tggttatgaa gaggaaaaat tggcagtaac
     3241 ctggccccac aaaccttcaa atgaacgaat caaattaaca accataggat gataatgcga
     3301 ttagtttttt agccttattt ctggggtaat taatcagcga agcgatgatt tttgatctat
     3361 taacagatat ataaatgcaa aaactgcata accactttaa ctaatacttt caacattttc
     3421 ggtttgtatt acttcttatt caaatgtaat aaaagtatca acaaaaaatt gttaatatac
     3481 ctctatactt taacgtcaag gagaaaaaac ccggatccat aatgagtgct gccactgcga
     3541 ctgctgcccc tgttcctcct cctgtaggga tatccaatct tcccaaccag cgttataaaa
     3601 ttgttaatga agaaggtggt acattcaccg tgatgttatg tggtgagagc ggattgggta
     3661 aaactacatt tattaatacc ttatttcaga ctgtcttgaa aagggcagat ggtcagcaac
     3721 accgccaaga acccatcaga aagactgtag aaatcgacat cacaagagct ttgctggaag
     3781 agaagcattt cgaattacgc gttaatgtca tcgacacccc aggttttgga gataacgtga
     3841 ataacaacaa agcctggcaa ccattggtgg attttattga tgatcaacac gattcttata
     3901 tgcgtcaaga gcagcaaccg taccgtacta agaaatttga cttgagagtt catgccgttc
     3961 tgtacttcat caggcccact ggacatggtt taaagccaat tgacattgag actatgaaga
     4021 gattgtcaac cagagccaat ctgattccag ttattgcgaa agctgataca ttgactgccc
     4081 aagaattaca acagttcaag tccagaatca ggcaagtaat agaagctcaa gaaattcgta
     4141 ttttcacgcc tccattagac gctgattcta aggaagatgc caaaagtggc agtaatccag
     4201 attctgccgc tgttgaacat gcaagacagt tgatagaagc aatgccattt gccatagtag
     4261 ggtctgaaaa aaaatttgac aatgggcaag gcactcaggt agttgcaagg aagtatcctt
     4321 ggggacttgt ggaaatcgaa aacgattctc attgtgattt ccgtaagctc agagccctgt
     4381 tactgaggac ttatctatta gatttgattt ctaccaccca agaaatgcat tacgaaacat
     4441 acagaagatt aagacttgag ggtcacgaga acaccgggga ggggaatgaa gatttcactc
     4501 ttccagcaat tgcaccagca aggaaattgt cacacaatcc tagatacaaa gaagaagaaa
     4561 atgcattgaa gaagtatttc actgaccaag tcaaagctga ggagcaaaga tttagacaat
     4621 ggcagcaaaa cattgttaac gaaagaatca gactgaacgg tgacttggaa gaaattcaag
     4681 gaaaggtaaa aaaactagaa gagcaggtca aaagcttgca agtaaaaaaa tcccatttaa
     4741 aatgatgatt aattaatgtc ttcctctttg tctcgtcaat ttcaacgcct atctctttgt
     4801 ctcttctttt gtcgcagcaa tacgcacata tctagagcgg ccgccaccgc ggtggagctc
     4861 caattcgccc tatagtgagt cgtattacaa ttcactggcc gtcgttttac aacgtcgtga
     4921 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     4981 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     5041 tggcgaatgg cgcgacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     5101 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     5161 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     5221 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     5281 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     5341 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     5401 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     5461 ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt cctgatgcgg
     5521 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tagggtaata actgatataa
     5581 ttaaattgaa gctctaattt gtgagtttag tatacatgca tttacttata atacagtttt
     5641 ttagttttgc tggccgcatc ttctcaaata tgcttcccag cctgcttttc tgtaacgttc
     5701 accctctacc ttagcatccc ttccctttgc aaatagtcct cttccaacaa taataatgtc
     5761 agatcctgta gagaccacat catccacggt tctatactgt tgacccaatg cgtctccctt
     5821 gtcatctaaa cccacaccgg gtgtcataat caaccaatcg taaccttcat ctcttccacc
     5881 catgtctctt tgagcaataa agccgataac aaaatctttg tcgctcttcg caatgtcaac
     5941 agtaccctta gtatattctc cagtagatag ggagcccttg catgacaatt ctgctaacat
     6001 caaaaggcct ctaggttcct ttgttacttc ttctgccgcc tgcttcaaac cgctaacaat
     6061 acctgggccc accacaccgt gtgcattcgt aatgtctgcc cattctgcta ttctgtatac
     6121 acccgcagag tactgcaatt tgactgtatt accaatgtca gcaaattttc tgtcttcgaa
     6181 gagtaaaaaa ttgtacttgg cggataatgc ctttagcggc ttaactgtgc cctccatgga
     6241 aaaatcagtc aagatatcca catgtgtttt tagtaaacaa attttgggac ctaatgcttc
     6301 aactaactcc agtaattcct tggtggtacg aacatccaat gaagcacaca agtttgtttg
     6361 cttttcgtgc atgatattaa atagcttggc agcaacagga ctaggatgag tagcagcacg
     6421 ttccttatat gtagctttcg acatgattta tcttcgtttc ctgcaggttt ttgttctgtg
     6481 cagttgggtt aagaatactg ggcaatttca tgtttcttca acactacata tgcgtatata
     6541 taccaatcta agtctgtgct ccttccttcg ttcttccttc tgttcggaga ttaccgaatc
     6601 aaaaaaattt caaagaaacc gaaatcaaaa aaaagaataa aaaaaaaatg atgaattgaa
     6661 ttgaaaagct gtggtatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     6721 ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     6781 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     6841 gtcatcaccg aaacgcgcga
//
