LOCUS       sequence-5-expor        6837 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 6837)
  AUTHORS   Liu JJ, Lindquist S
  TITLE     Oligopeptide-repeat expansions modulate 'protein-only' inheritance 
            in yeast.
  JOURNAL   Nature 1999 Aug 5;400(6744):573-6.
  PUBMED    10448860
REFERENCE   2  (bases 1 to 6837)
  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: "Nature"; 
            date: "1999-08-5"; volume: "400(6744)"; pages: "573-6"
COMMENT     Sequence Label: NM R2E2 GFP
FEATURES             Location/Qualifiers
     source          1..6837
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        196..416
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             417..1220
                     /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"
     rep_origin      complement(1351..1806)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1488..1509)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1700..1719
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1936..1958
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1950..1967
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1951..1967
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1974..1993
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1974..1992
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             complement(2010..2726)
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MASKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(2050..2070)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     2665..2693
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     promoter        complement(3599..3833)
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
     primer_bind     complement(4017..4033)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4018..4034)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(4063..4081)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4102..4118)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4102..4118)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4115..4137)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4126..4142
                     /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(4150..4156,4157..4174,4175..4180))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4333..4350)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4504..5092)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4584..4603)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5263..6054,6055..6123))
                     /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     5886..5905
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6124..6228)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6265..6768
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     6810..6828
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataccac agcttttcaa ttcaattcat catttttttt ttattctttt ttttgatttc
      241 ggtttctttg aaattttttt gattcggtaa tctccgaaca gaaggaagaa cgaaggaagg
      301 agcacagact tagattggta tatatacgca tatgtagtgt tgaagaaaca tgaaattgcc
      361 cagtattctt aacccaactg cacagaacaa aaacctgcag gaaacgaaga taaatcatgt
      421 cgaaagctac atataaggaa cgtgctgcta ctcatcctag tcctgttgct gccaagctat
      481 ttaatatcat gcacgaaaag caaacaaact tgtgtgcttc attggatgtt cgtaccacca
      541 aggaattact ggagttagtt gaagcattag gtcccaaaat ttgtttacta aaaacacatg
      601 tggatatctt gactgatttt tccatggagg gcacagttaa gccgctaaag gcattatccg
      661 ccaagtacaa ttttttactc ttcgaagaca gaaaatttgc tgacattggt aatacagtca
      721 aattgcagta ctctgcgggt gtatacagaa tagcagaatg ggcagacatt acgaatgcac
      781 acggtgtggt gggcccaggt attgttagcg gtttgaagca ggcggcagaa gaagtaacaa
      841 aggaacctag aggccttttg atgttagcag aattgtcatg caagggctcc ctatctactg
      901 gagaatatac taagggtact gttgacattg cgaagagcga caaagatttt gttatcggct
      961 ttattgctca aagagacatg ggtggaagag atgaaggtta cgattggttg attatgacac
     1021 ccggtgtggg tttagatgac aagggagacg cattgggtca acagtataga accgtggatg
     1081 atgtggtctc tacaggatct gacattatta ttgttggaag aggactattt gcaaagggaa
     1141 gggatgctaa ggtagagggt gaacgttaca gaaaagcagg ctgggaagca tatttgagaa
     1201 gatgcggcca gcaaaactaa aaaactgtat tataagtaaa tgcatgtata ctaaactcac
     1261 aaattagagc ttcaatttaa ttatatcagt tattacccta tgcggtgtga aataccgcac
     1321 agatgcgtaa ggagaaaata ccgcatcagg aaattgtaaa cgttaatatt ttgttaaaat
     1381 tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa
     1441 tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca
     1501 agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg
     1561 gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta
     1621 aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg
     1681 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa
     1741 gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg
     1801 gcgcgtcgcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     1861 cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg
     1921 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgaa ttgtaatacg
     1981 actcactata gggcgaattg gagctctcat ttgtatagtt catccatgcc atgtgtaatc
     2041 ccagcagctg ttacaaactc aagaaggacc atgtggtctc tcttttcgtt gggatctttc
     2101 gaaagggcag attgtgtgga caggtaatgg ttgtctggta aaaggacagg gccatcgcca
     2161 attggagtat tttgttgata atggtctgct agttgaacgc ttccatcttc aatgttgtgt
     2221 ctaattttga agttagcttt gattccattc ttttgtttgt ctgccatgat gtatacattg
     2281 tgtgagttat agttgtattc caatttgtgc ccaagaatgt ttccatcttc tttaaaatca
     2341 atacctttta actcgattct attaacaagg gtatcacctt caaacttgac ttcagcacgt
     2401 gtcttgtagt tcccgtcatc tttgaaaaat atagttcttt cctgtacata accttcgggc
     2461 atggcactct tgaaaaagtc atgccgtttc atatgatccg ggtatcttga aaagcactga
     2521 acaccataag tgaaagtagt gacaagtgtt ggccatggaa caggtagttt tccagtagtg
     2581 caaataaatt taagggtaag ttttccgtat gttgcatcac cttcaccctc tccactgaca
     2641 gaaaatttgt gcccattaac atcaccatct aattcaacaa gaattgggac aactccagtg
     2701 aagagttctt ctcctttgct agccatgagc tcatcgttaa caacttcgtc atccacttct
     2761 tcttcctgtt ccttgatcaa ggcatcagca ctggtaacat tggcattgtt ggtattatgt
     2821 gttgattcag agattttaag gtcttctacc tttggaagtt ccgatttttc ctccgtcttt
     2881 tcttcagtct ggactggttt ctcctccttt ttaactggtt cttcgacctt tgttggctct
     2941 ttagttggtt ctttggtttc agcagacttc tcttcctctt tcttatcaga ttcggcaggt
     3001 tttgtgccaa ccttcttggt agcattggcc aacttgatac cggaactgga gacaagcttc
     3061 aaagtcttct ttggtttggg agcggcctgc ttttgttgct tttgaaagtc gttcaaagac
     3121 ataccttgag actgtggttg gaaaccagct tgatatcctt gcaaattgtt attgtagttg
     3181 aagtttttgt aatttccacg gccaccttgt ggattgaatt gctgctgata accgccttga
     3241 ggattgtact gttgatagcc tccttgagga ttatactgtt gctggtaacc ggcgtcggga
     3301 ttgtactgtt gatagccacc ttggggattg tactgttgat agccaccttg gggattgtac
     3361 tgttgatagc caccttgttg gtacccagaa taaccttggt aattttggta gtacccacct
     3421 gcaggttggg cttgagcatt gtaagcttga taaccttggt atctgttgtt accttgttgt
     3481 tggttaccgt tctggctgta ttgctggtag ttttgctgat tgttgccttg gtttgaatcc
     3541 gacatggatc ctgattgatt gattgattgt acagtttgtt tttcttaata tctatttcga
     3601 tgacttctat atgatattgc actaacaaga agatattata atgcaattga tacaagacaa
     3661 ggagttattt gcttctcttt tatatgattc tgacaatcca tattgcgttg gtagtctttt
     3721 ttgctggaac ggttcagcgg aaaagacgca tcgctctttt tgcttctaga agaaatgcca
     3781 gcaaaagaat ctcttgacag tgactgacag caaaaatgtc tttttctaac tagtaacaag
     3841 gctaagatat cagcctgaaa taaagggtgg tgaagtaata attaaatcat ccgtataaac
     3901 ctatacacat atatgaggaa aaataataca aaagtgtttt aaatacagat acatacatga
     3961 acatatgcac gtatagcgcc caaatgtcgg taatgggaat tcgatatcaa gcttatcgat
     4021 accgtcgacc tcgagggggg gcccggtacc cagcttttgt tccctttagt gagggttaat
     4081 tccgagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     4141 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     4201 gaggtaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     4261 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4321 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4381 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4441 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4501 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4561 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4621 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4681 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4741 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4801 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4861 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4921 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     4981 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     5041 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     5101 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     5161 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     5221 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     5281 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     5341 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     5401 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     5461 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     5521 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5581 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5641 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5701 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5761 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     5821 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     5881 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5941 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     6001 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     6061 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     6121 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     6181 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     6241 aaaagtgcca cctgggtcct tttcatcacg tgctataaaa ataattataa tttaaatttt
     6301 ttaatataaa tatataaatt aaaaatagaa agtaaaaaaa gaaattaaag aaaaaatagt
     6361 ttttgttttc cgaagatgta aaagactcta gggggatcgc caacaaatac taccttttat
     6421 cttgctcttc ctgctctcag gtattaatgc cgaattgttt catcttgtct gtgtagaaga
     6481 ccacacacga aaatcctgtg attttacatt ttacttatcg ttaatcgaat gtatatctat
     6541 ttaatctgct tttcttgtct aataaatata tatgtaaagt acgctttttg ttgaaatttt
     6601 ttaaaccttt gtttattttt ttttcttcat tccgtaactc ttctaccttc tttatttact
     6661 ttctaaaatc caaatacaaa acataaaaat aaataaacac agagtaaatt cccaaattat
     6721 tccatcatta aaagatacga ggcgcgtgta agttacaggc aagcgatccg tcctaagaaa
     6781 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtc
//
