LOCUS       sequence-164746-        6462 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 6462)
  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 6462)
  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: CDC12(WT)
FEATURES             Location/Qualifiers
     source          1..6462
                     /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    69..572
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        609..713
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(714..782,783..1574)
                     /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(932..951)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1745..2333
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2234..2253
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2487..2504
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2657..2662,2663..2680,2681..2687)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2695..2711
                     /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     2700..2722
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2719..2735
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2719..2735
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2756..2774
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     2802..2818
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2803..2819
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4470..4489)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4471..4489)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4496..4513)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4496..4512)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4505..4527)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4657..5112
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4744..4763)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4954..4975
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5243..6046)
                     /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(6047..6267)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(6293..6312)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6412..6434
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgga tcgcttgcct gtaacttaca cgcgcctcgt atcttttaat gatggaataa
      121 tttgggaatt tactctgtgt ttatttattt ttatgttttg tatttggatt ttagaaagta
      181 aataaagaag gtagaagagt tacggaatga agaaaaaaaa ataaacaaag gtttaaaaaa
      241 tttcaacaaa aagcgtactt tacatatata tttattagac aagaaaagca gattaaatag
      301 atatacattc gattaacgat aagtaaaatg taaaatcaca ggattttcgt gtgtggtctt
      361 ctacacagac aagatgaaac aattcggcat taatacctga gagcaggaag agcaagataa
      421 aaggtagtat ttgttggcga tccccctaga gtcttttaca tcttcggaaa acaaaaacta
      481 ttttttcttt aatttctttt tttactttct atttttaatt tatatattta tattaaaaaa
      541 tttaaattat aattattttt atagcacgtg atgaaaagga cccaggtggc acttttcggg
      601 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
      661 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
      721 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
      781 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
      841 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
      901 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
      961 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     1021 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     1081 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     1141 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     1201 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     1261 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     1321 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     1381 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     1441 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     1501 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     1561 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     1621 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     1681 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     1741 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     1801 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     1861 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     1921 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     1981 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2041 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     2101 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     2161 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     2221 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     2281 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     2341 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     2401 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     2461 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     2521 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     2581 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tacctcactc
     2641 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     2701 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag ctcgaaatta
     2761 accctcacta aagggaacaa aagctggtac cgggcccccc ctcgaggtcg acggtatcga
     2821 taagcttgat atcgaattcc tgcagcccgg gggatccgat gcagtatcac gattagcaat
     2881 tcagctatga gttatgttgc tctttgtttt gtttatggaa attgtcctat ggtaagtctc
     2941 ttttttttgc aatcgtgatt acagaaaaaa aacagggcgc tggaaaagtg aagaatccga
     3001 aatttttttc gaaatcacca ttgtttgttt tgagtagatc aaagtcttga aaggtgcagc
     3061 aagatatagg atcttgacct gaagagtatt gataacgaac tacatcacat attgtatcaa
     3121 ataatgagtg ctgccactgc gactgctgcc cctgttcctc ctcctgtagg gatatccaat
     3181 cttcccaacc agcgttataa aattgttaat gaagaaggtg gtacattcac cgtgatgtta
     3241 tgtggtgaga gcggattggg taaaactaca tttattaata ccttatttca gactgtcttg
     3301 aaaagggcag atggtcagca acaccgccaa gaacccatca gaaagactgt agaaatcgac
     3361 atcacaagag ctttgctgga agagaagcat ttcgaattac gcgttaatgt catcgacacc
     3421 ccaggttttg gagataacgt gaataacaac aaagcctggc aaccattggt ggattttatt
     3481 gatgatcaac acgattctta tatgcgtcaa gagcagcaac cgtaccgtac taagaaattt
     3541 gacttgagag ttcatgccgt tctgtacttc atcaggccca ctggacatgg tttaaagcca
     3601 attgacattg agactatgaa gagattgtca accagagcca atctgattcc agttattgcg
     3661 aaagctgata cattgactgc ccaagaatta caacagttca agtccagaat caggcaagta
     3721 atagaagctc aagaaattcg tattttcacg cctccattag acgctgattc taaggaagat
     3781 gccaaaagtg gcagtaatcc agattctgcc gctgttgaac atgcaagaca gttgatagaa
     3841 gcaatgccat ttgccatagt agggtctgaa aaaaaatttg acaatgggca aggcactcag
     3901 gtagttgcaa ggaagtatcc ttggggactt gtggaaatcg aaaacgattc tcattgtgat
     3961 ttccgtaagc tcagagccct gttactgagg acttatctat tagatttgat ttctaccacc
     4021 caagaaatgc attacgaaac atacagaaga ttaagacttg agggtcacga gaacaccggg
     4081 gaggggaatg aagatttcac tcttccagca attgcaccag caaggaaatt gtcacacaat
     4141 cctagataca aagaagaaga aaatgcattg aagaagtatt tcactgacca agtcaaagct
     4201 gaggagcaaa gatttagaca atgggagcaa aacattgtta acgaaagaat cagactgaac
     4261 ggtgacttgg aagaaattca aggaaaggta aaaaaactag aagagcaggt caaaagcttg
     4321 caagtaaaaa aatcccattt aaaatgatga ttaattaatg tcttcctctt tgtctcgtca
     4381 atttcaacgc ctatctcttt gtctcttctt ttgtcgcagc aatacgcaca tatctagagc
     4441 ggccgccacc gcggtggagc tccaattcgc cctatagtga gtcgtattac aattcactgg
     4501 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
     4561 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
     4621 cccaacagtt gcgcagcctg aatggcgaat ggcgcgacgc gccctgtagc ggcgcattaa
     4681 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     4741 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     4801 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4861 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4921 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4981 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     5041 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     5101 cgtttacaat ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     5161 catagggtaa taactgatat aattaaattg aagctctaat ttgtgagttt agtatacatg
     5221 catttactta taatacagtt ttttagtttt gctggccgca tcttctcaaa tatgcttccc
     5281 agcctgcttt tctgtaacgt tcaccctcta ccttagcatc ccttcccttt gcaaatagtc
     5341 ctcttccaac aataataatg tcagatcctg tagagaccac atcatccacg gttctatact
     5401 gttgacccaa tgcgtctccc ttgtcatcta aacccacacc gggtgtcata atcaaccaat
     5461 cgtaaccttc atctcttcca cccatgtctc tttgagcaat aaagccgata acaaaatctt
     5521 tgtcgctctt cgcaatgtca acagtaccct tagtatattc tccagtagat agggagccct
     5581 tgcatgacaa ttctgctaac atcaaaaggc ctctaggttc ctttgttact tcttctgccg
     5641 cctgcttcaa accgctaaca atacctgggc ccaccacacc gtgtgcattc gtaatgtctg
     5701 cccattctgc tattctgtat acacccgcag agtactgcaa tttgactgta ttaccaatgt
     5761 cagcaaattt tctgtcttcg aagagtaaaa aattgtactt ggcggataat gcctttagcg
     5821 gcttaactgt gccctccatg gaaaaatcag tcaagatatc cacatgtgtt tttagtaaac
     5881 aaattttggg acctaatgct tcaactaact ccagtaattc cttggtggta cgaacatcca
     5941 atgaagcaca caagtttgtt tgcttttcgt gcatgatatt aaatagcttg gcagcaacag
     6001 gactaggatg agtagcagca cgttccttat atgtagcttt cgacatgatt tatcttcgtt
     6061 tcctgcaggt ttttgttctg tgcagttggg ttaagaatac tgggcaattt catgtttctt
     6121 caacactaca tatgcgtata tataccaatc taagtctgtg ctccttcctt cgttcttcct
     6181 tctgttcgga gattaccgaa tcaaaaaaat ttcaaggaaa ccgaaatcaa aaaaaagaat
     6241 aaaaaaaaaa tgatgaattg aattgaaaag ctgtggtatg gtgcactctc agtacaatct
     6301 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     6361 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     6421 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc ga
//
