LOCUS       sequence-243811-        6527 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 6527)
  AUTHORS   Duennwald ML, Jagadish S, Muchowski PJ, Lindquist S
  TITLE     Flanking sequences profoundly alter polyglutamine toxicity in yeast.
  JOURNAL   Proc Natl Acad Sci U S A. 2006 Jul 18. 103(29):11045-50.
  PUBMED    16832050
REFERENCE   2  (bases 1 to 6527)
  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: "Proc Natl 
            Acad Sci U S A. 2006 Jul 18. 103(29):11045-50."
COMMENT     Sequence Label: GAL FLAG103Q+ProGFPp303
FEATURES             Location/Qualifiers
     source          1..6527
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..442
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    1..118
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     413..436
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             469..492
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     1079..1100
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             1155..1865
                     /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="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKAH
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(1182..1210)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1805..1825
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      1875..2122
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1891..1909)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2139..2155)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2140..2156)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(2184..2202)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2223..2239)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2223..2239)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2236..2258)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2247..2263
                     /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(2271..2277,2278..2295,2296..2301))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2316..2337
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2454..2471)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2625..3213)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2705..2724)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3384..4175,4176..4244))
                     /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     4007..4026
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4245..4349)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4417..4435
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4473..4495)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4595..4614
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4761..4948
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             4949..5611
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(5871..6326)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6008..6029)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6220..6239
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6452..6474
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6466..6483
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6467..6483
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6490..6509
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6490..6508
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 cggattagaa gccgccgagc gggtgacagc cctccgaagg aagactctcc tccgtgcgtc
       61 ctcgtcttca ccggtcgcgt tcctgaaacg cagatgtgcc tcgcgccgca ctgctccgaa
      121 caataaagat tctacaatac tagcttttat ggttatgaag aggaaaaatt ggcagtaacc
      181 tggccccaca aaccttcaaa tgaacgaatc aaattaacaa ccataggatg ataatgcgat
      241 tagtttttta gccttatttc tggggtaatt aatcagcgaa gcgatgattt ttgatctatt
      301 aacagatata taaatgcaaa aactgcataa ccactttaac taatactttc aacattttcg
      361 gtttgtatta cttcttattc aaatgtaata aaagtatcaa caaaaaattg ttaatatacc
      421 tctatacttt aacgtcaagg agaaaaaacc ccggattcta gagccatgga ctacaaggac
      481 gacgatgaca agctggcgac cctggaaaag ctgatgaagg ccttcgagtc cctcaaaagc
      541 ttccaacagc agcaacagca acaacagcag caacagcaac aacagcagca acagcaacaa
      601 cagcagcaac agcaacaaca gcagcaacag caacaacagc agcaacagca acaacagcag
      661 caacagcaac aacagcagca acagcaacaa cagcagcaac agcaacaaca gcagcaacag
      721 caacaacagc agcaacagca acaacagcag caacagcaac aacagcagca acagcaacaa
      781 cagcagcaac agcaacaaca gcagcaacag caacaacagc agcaacagca acaaccgcca
      841 ccacctcccc ctccaccccc acctcctcaa cttcctcaac ctcctccaca ggcacagcct
      901 ctgctgcctc agccacaacc tcctccacct ccacctccac ctcctccagg cccagctgtg
      961 gctgaggagc ctctgcaccg acctggatcc aggtcggtgc agatgaactt cagggtcagc
     1021 ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg
     1081 tcgccgtcca gctcgaccag gatgggcacc accccggtga acagctcctc gcccttgctc
     1141 accagggatc cctgagtaaa ggagaagaac ttttcactgg agttgtccca attcttgttg
     1201 aattagatgg tgatgttaat gggcacaaat tttctgtcag tggagagggt gaaggtgatg
     1261 caacatacgg aaaacttacc cttaaattta tttgcactac tggaaaacta cctgttccat
     1321 ggccaacact tgtcactact ttgacttggg gtgttcaatg cttttcaaga tacccagatc
     1381 atatgaaacg gcatgacttt ttcaagagtg ccatgcccga aggttatgta caggaaagaa
     1441 ctatattttt caaagatgac gggaactaca agacacgtgc tgaagtcaag tttgaaggtg
     1501 atacccttgt taatagaatc gagttaaaag gtattgattt taaagaagat ggaaacattc
     1561 ttggacacaa attggaatac aactatattt cacacaatgt atacatcact gcagacaaac
     1621 aaaagaatgg aatcaaagct catttcaaaa ttagacacaa cattgaagat ggaagcgttc
     1681 aactagcaga ccattatcaa caaaatactc caattggcga tggccctgtc cttttaccag
     1741 acaaccatta cctgtccaca caatctgccc tttcgaaaga tcccaacgaa aagagagacc
     1801 acatggtcct tcttgagttt gtaacagctg ctgggattac acatggcatg gatgaactat
     1861 acaaatagct cgagtcatgt aattagttat gtcacgctta cattcacgcc ctccccccac
     1921 atccgctcta accgaaaagg aaggagttag acaacctgaa gtctaggtcc ctatttattt
     1981 ttttatagtt atgttagtat taagaacgtt atttatattt caaatttttc ttttttttct
     2041 gtacagacgc gtgtacgcat gtaacattat actgaaaacc ttgcttgaga aggttttggg
     2101 acgctcgaag gctttaattt gcggccgatc aagcttatcg ataccgtcga cctcgagggg
     2161 gggcccggta ccagcttttg ttccctttag tgagggttaa tttcgagctt ggcgtaatca
     2221 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     2281 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     2341 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     2401 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     2461 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     2521 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     2581 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     2641 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2701 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2761 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2821 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2881 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2941 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     3001 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     3061 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     3121 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     3181 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     3241 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     3301 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     3361 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     3421 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     3481 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     3541 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     3601 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     3661 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     3721 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     3781 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     3841 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     3901 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     3961 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     4021 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     4081 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     4141 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     4201 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     4261 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     4321 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     4381 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     4441 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     4501 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     4561 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga
     4621 gtgcaccata aattcccgtt ttaagagctt ggtgagcgct aggagtcact gccaggtatc
     4681 gtttgaacac ggcattagtc agggaagtca taacacagtc ctttcccgca attttctttt
     4741 tctattactc ttggcctcct ctagtacact ctatattttt ttatgcctcg gtaatgattt
     4801 tcattttttt ttttccacct agcggatgac tctttttttt tcttagcgat tggcattatc
     4861 acataatgaa ttatacatta tataaagtaa tgtgatttct tcgaagaata tactaaaaaa
     4921 tgagcaggca agataaacga aggcaaagat gacagagcag aaagccctag taaagcgtat
     4981 tacaaatgaa accaagattc agattgcgat ctctttaaag ggtggtcccc tagcgataga
     5041 gcactcgatc ttcccagaaa aagaggcaga agcagtagca gaacaggcca cacaatcgca
     5101 agtgattaac gtccacacag gtatagggtt tctggaccat atgatacatg ctctggccaa
     5161 gcattccggc tggtcgctaa tcgttgagtg cattggtgac ttacacatag acgaccatca
     5221 caccactgaa gactgcggga ttgctctcgg tcaagctttt aaagaggccc taggggccgt
     5281 gcgtggagta aaaaggtttg gatcaggatt tgcgcctttg gatgaggcac tttccagagc
     5341 ggtggtagat ctttcgaaca ggccgtacgc agttgtcgaa cttggtttgc aaagggagaa
     5401 agtaggagat ctctcttgcg agatgatccc gcattttctt gaaagctttg cagaggctag
     5461 cagaattacc ctccacgttg attgtctgcg aggcaagaat gatcatcacc gtagtgagag
     5521 tgcgttcaag gctcttgcgg ttgccataag agaagccacc tcgcccaatg gtaccaacga
     5581 tgttccctcc accaaaggtg ttcttatgta gtgacaccga ttatttaaag ctgcagcata
     5641 cgatatatat acatgtgtat atatgtatac ctatgaatgt cagtaagtat gtatacgaac
     5701 agtatgatac tgaagatgac aaggtaatgc atcattctat acgtgtcatt ctgaacgagg
     5761 cgcgctttcc ttttttcttt ttgctttttc tttttttttc tcttgaactc gacggatcta
     5821 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg aaattgtaag
     5881 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     5941 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     6001 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     6061 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     6121 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     6181 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     6241 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     6301 gcttaatgcg ccgctacagg gcgcgtccat tcgccattca ggctgcgcaa ctgttgggaa
     6361 gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca
     6421 aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc
     6481 agtgaattgt aatacgactc actatagggc gaattggagc tctagta
//
