LOCUS       sequence-18-expo        6669 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 6669)
  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 6669)
  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  R(delta) 2-5 GFP
FEATURES             Location/Qualifiers
     source          1..6669
                     /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(3431..3665)
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
     primer_bind     complement(3849..3865)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3850..3866)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(3895..3913)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3934..3950)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3934..3950)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3947..3969)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3958..3974
                     /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(3982..3988,3989..4006,4007..4012))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4165..4182)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4336..4924)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4416..4435)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5095..5886,5887..5955))
                     /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     5718..5737
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5956..6060)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6097..6600
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     6642..6660
                     /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 gggtacccag aataaccttg gtaattttgg
     3241 tagtacccac ctgcaggttg ggcttgagca ttgtaagctt gataaccttg gtatctgttg
     3301 ttaccttgtt gttggttacc gttctggctg tattgctggt agttttgctg attgttgcct
     3361 tggtttgaat ccgacatgga tcctgattga ttgattgatt gtacagtttg tttttcttaa
     3421 tatctatttc gatgacttct atatgatatt gcactaacaa gaagatatta taatgcaatt
     3481 gatacaagac aaggagttat ttgcttctct tttatatgat tctgacaatc catattgcgt
     3541 tggtagtctt ttttgctgga acggttcagc ggaaaagacg catcgctctt tttgcttcta
     3601 gaagaaatgc cagcaaaaga atctcttgac agtgactgac agcaaaaatg tctttttcta
     3661 actagtaaca aggctaagat atcagcctga aataaagggt ggtgaagtaa taattaaatc
     3721 atccgtataa acctatacac atatatgagg aaaaataata caaaagtgtt ttaaatacag
     3781 atacatacat gaacatatgc acgtatagcg cccaaatgtc ggtaatggga attcgatatc
     3841 aagcttatcg ataccgtcga cctcgagggg gggcccggta cccagctttt gttcccttta
     3901 gtgagggtta attccgagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     3961 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     4021 tgcctaatga gtgaggtaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4081 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4141 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4201 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4261 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4321 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4381 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4441 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     4501 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     4561 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     4621 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     4681 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     4741 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     4801 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     4861 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     4921 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     4981 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5041 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5101 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5161 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5221 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     5281 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     5341 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     5401 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     5461 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     5521 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     5581 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     5641 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     5701 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     5761 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     5821 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     5881 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     5941 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6001 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6061 cacatttccc cgaaaagtgc cacctgggtc cttttcatca cgtgctataa aaataattat
     6121 aatttaaatt ttttaatata aatatataaa ttaaaaatag aaagtaaaaa aagaaattaa
     6181 agaaaaaata gtttttgttt tccgaagatg taaaagactc tagggggatc gccaacaaat
     6241 actacctttt atcttgctct tcctgctctc aggtattaat gccgaattgt ttcatcttgt
     6301 ctgtgtagaa gaccacacac gaaaatcctg tgattttaca ttttacttat cgttaatcga
     6361 atgtatatct atttaatctg cttttcttgt ctaataaata tatatgtaaa gtacgctttt
     6421 tgttgaaatt ttttaaacct ttgtttattt ttttttcttc attccgtaac tcttctacct
     6481 tctttattta ctttctaaaa tccaaataca aaacataaaa ataaataaac acagagtaaa
     6541 ttcccaaatt attccatcat taaaagatac gaggcgcgtg taagttacag gcaagcgatc
     6601 cgtcctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     6661 cctttcgtc
//
