LOCUS       sequence-450960-        8877 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for cytoplasmic aggregates under LEU2 selection
            marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8877)
  TITLE     Budding yeast expression plasmid for induced production of 
            fluorescently tagged cytoplasmic protein aggregates
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8877)
  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: 
            "Unpublished"
COMMENT     Sequence Label: CUP1-∆ssCPY*-mScarletI in pRS315
FEATURES             Location/Qualifiers
     source          1..8877
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     274..293
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     527..544
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    661..682
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(697..702,703..720,721..727)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    735..751
                     /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     740..762
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     759..775
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     759..775
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        796..814
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     980..998
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(3417..3651)
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
     primer_bind     complement(3814..3833)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3815..3833)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3840..3857)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3840..3856)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3849..3871)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3997..4452
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4084..4103)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4294..4315
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4753..5160
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             5161..6255
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     primer_bind     complement(6748..6767)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6867..6889
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6927..6945)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    6986..7489
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        7526..7630
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7631..7699,7700..8491)
                     /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(7849..7868)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(8662..8877,1..373)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
       61 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
      121 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
      181 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
      241 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
      301 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
      361 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
      421 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
      481 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
      541 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
      601 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
      661 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
      721 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga
      781 ttacgccaag ctcggaatta accctcacta aagggaacaa aagctgggta ccgtacacgc
      841 gtttgtacag aaaaaaaaga aaaatttgaa atataaataa cgttcttaat actaacataa
      901 ctataaaaaa ataaataggg acctagactt caggttgtct aactccttcc ttttcggtta
      961 gagcggatgt ggggggaggg cgtgaatgta agcgtgacat aactaattac atgatatcga
     1021 caaaggaaaa ggggcctgtt tactcacagg cttttttcaa gtaggtaatt aagtcgtttc
     1081 tgtctttttc cttcttcaac ccggtaccgg atccagatat tctatggcaa agcttttatt
     1141 tatacagctc atccatacca cccgtggaat ggcggccttc acttctctca tactgctcca
     1201 ccactgtgta atcttcgtta tggctggtaa tgtccagctt acgatccacg ttatacgccc
     1261 ctggcatttg aacaggtttt ttcgctttgt atgtggtctt aaagtcagcg aggtatctgc
     1321 ctccatcctt gagacggagt gccattttga tatctccctt taaaacaccg tcctctggat
     1381 agagacgctc cgttgatgcc tcccaaccca tcgttttttt ctgcataaca ggcccgtcag
     1441 gaggaaaatt tgtccctctg agcttgacct tataaatcag cgtgccgtct tcaagactgg
     1501 tatcctgcgt cacggtgaca gctcctccgt cttcaaagtt cattactctt tcccacttga
     1561 acccttctgg gaagctctgc ttgtaatagt caggtatatc agcagggtgt ttgatgaaag
     1621 ctctggaccc atacatgaat tgtgggctta aaatatccca actaaatgga agtgggccac
     1681 ctttggtaac tttcaattta gcagtctgcg taccttcgta aggccgccct tcgccctctc
     1741 cttctatctc gaactcatgg ccgttcatag acccctccat gtgaactttg aatctcatga
     1801 actctttgat tacggcctct cctttggaga ccatctcgag taaggagaaa ccaccgtgga
     1861 tccattcgtt aaccatactt aaggcgtttt cagggacgtc aaatggaacc atgtggccac
     1921 cattgaagac tctcaaatag gtgaagtgct tgtaggattt gacttcacca gcgacctcgt
     1981 cggtgataga agcagtccag ttacgtactt tttggcttgc aaattcttcg tcgtacttcc
     2041 atggcaagac atccgtccac gccttattac ccaaccagtt acagatgaaa tctttatcgc
     2101 ctgcatatac cagaatgggt aggtcttgat tcaaaagatc tgttacggcg gtgtggtaag
     2161 gcttcatcca atcacccgca aacaggaaat ttctgttgat atcgaagtta caggattcgt
     2221 agtggtcaac ctccgcaccg acagcttctt tgacgtagtc ctggtttaag tagtcgtcga
     2281 tatcttgtaa cgttgggtag cacaaattgc caccttcaca atccttcctg atatcgtaaa
     2341 cgtttctgcc ggtacgttgg taaggagcca attgggcgtt attacaataa atggtagctg
     2401 gaacacagga ccagacggat tgcgagtcat agcacgactc gatcaagccc aaacaacgtt
     2461 ccaaagagtc ttccatagca gagcattcct ccgagggcaa aacagatggt tcgccacctt
     2521 caccacaggc cattggttcg taatagttat actgagtcaa tgggtcagtg aggccatttc
     2581 cgatcaagac ggaggttaag ttgaagtttc tgtccttgtg agacaaaatt tcagaggcaa
     2641 aaacagggat gtaatggccg gcgtaggatt ccctagcgat gtggaaatct tggcccttgt
     2701 tgacgtattc agggaactga tcgaagaaca actccaagaa gttatagaca tccttaccag
     2761 cggcgacagt gttggaaaca cctgaggacc cggaatacga gaacccaacg ttgacaggct
     2821 ggtcaaggaa gatcacggtg gcattgctgt tccaagagta agggttcccg atgggtttca
     2881 aatcaggtcc aatggatgag ggtcctaatt caaagaacag cccggttagt gaagaacaac
     2941 ctggaccccc gttcaaccaa aggatgaccg gatcctttgc aggatcgttt ctactttcaa
     3001 aagtccaaaa gaagaaatgc ttgtcctcgt cttccacatc caagtaaccc gtgtactgtg
     3061 tgacatttgg gtcaatgccc aggattttag ggtccttaat cttgttgaca cgaagctgat
     3121 agttttcaat tgcgtcattc ttgaccacaa agtcccagtc tttcttcgtt ttgattgctt
     3181 cagggaattt tggcttagtg gaagtttcaa ggctcataac ctggtttggg tacaaatgct
     3241 ctatttgggc ccaagcgtcc aatacattgg agtccaactc cttcaagaga tgatccaggt
     3301 cgaggtccaa accaaatttt tccgcagctt gcagcaaaac gtccttatct agacccaacg
     3361 gtctttgcaa tgagatcatg aattcgttac agtttgtttt tcttaatatc tatttcgatg
     3421 acttctatat gatattgcac taacaagaag atattataat gcaattgaca caagacaagg
     3481 agttatttgc ttctctttta tatgattctg acaatccata ttgcgttggt agtctttttt
     3541 gctggaacgg ttcagcggaa aagacgcatc gctctttttg cttctagaag aaatgccagc
     3601 aaaagaatct cttgacagtg actgacagca aaaatgtctt tttctaacta gtaacaaggc
     3661 taagatatca gcctgaaata aagggtggtg aagtaataat taaatcatcc gtataaacct
     3721 atacacatat atgaggaaaa ataatacaaa agtgttttaa atacagatac atacatgaac
     3781 atgcggccgc caccgcggtg gagctccaat tcgccctata gtgagtcgta ttacaattca
     3841 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc
     3901 cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc
     3961 ccttcccaac agttgcgcag cctgaatggc gaatggacgc gccctgtagc ggcgcattaa
     4021 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     4081 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     4141 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4201 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4261 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4321 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     4381 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattag
     4441 cgcttacaat ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     4501 catatcgacc ggtcgaggag aacttctagt atatctacat acctaatatt attgccttat
     4561 taaaaatgga atcccaacaa ttacatcaaa atccacattc tcttcaaaat caattgtcct
     4621 gtacttcctt gttcatgtgt gttcaaaaac gttatattta taggataatt atactctatt
     4681 tctcaacaag taattggttg tttggccgag cggtctaagg cgcctgattc aagaaatatc
     4741 ttgaccgcag ttaactgtgg gaatactcag gtatcgtaag atgcaagagt tcgaatctct
     4801 tagcaaccat tatttttttc ctcaacataa cgagaacaca caggggcgct atcgcacaga
     4861 atcaaattcg atgactggaa attttttgtt aatttcagag gtcgcctgac gcatatacct
     4921 ttttcaactg aaaaattggg agaaaaagga aaggtgagag cgccggaacc ggcttttcat
     4981 atagaataga gaagcgttca tgactaaatg cttgcatcac aatacttgaa gttgacaata
     5041 ttatttaagg acctattgtt ttttccaata ggtggttagc aatcgtctta ctttctaact
     5101 tttcttacct tttacatttc agcaatatat atatatatat ttcaaggata taccattcta
     5161 atgtctgccc ctaagaagat cgtcgttttg ccaggtgacc acgttggtca agaaatcaca
     5221 gccgaagcca ttaaggttct taaagctatt tctgatgttc gttccaatgt caagttcgat
     5281 ttcgaaaatc atttaattgg tggtgctgct atcgatgcta caggtgttcc acttccagat
     5341 gaggcgctgg aagcctccaa gaaggctgat gccgttttgt taggtgctgt gggtggtcct
     5401 aaatggggta ccggtagtgt tagacctgaa caaggtttac taaaaatccg taaagaactt
     5461 caattgtacg ccaacttaag accatgtaac tttgcatccg actctctttt agacttatct
     5521 ccaatcaagc cacaatttgc taaaggtact gacttcgttg ttgtcagaga attagtggga
     5581 ggtatttact ttggtaagag aaaggaagac gatggtgatg gtgtcgcttg ggatagtgaa
     5641 caatacaccg ttccagaagt gcaaagaatc acaagaatgg ccgctttcat ggccctacaa
     5701 catgagccac cattgcctat ttggtccttg gataaagcta atgttttggc ctcttcaaga
     5761 ttatggagaa aaactgtgga ggaaaccatc aagaacgaat tccctacatt gaaggttcaa
     5821 catcaattga ttgattctgc cgccatgatc ctagttaaga acccaaccca cctaaatggt
     5881 attataatca ccagcaacat gtttggtgat atcatctccg atgaagcctc cgttatccca
     5941 ggttccttgg gtttgttgcc atctgcgtcc ttggcctctt tgccagacaa gaacaccgca
     6001 tttggtttgt acgaaccatg ccacggttct gctccagatt tgccaaagaa taaggtcaac
     6061 cctatcgcca ctatcttgtc tgctgcaatg atgttgaaat tgtcattgaa cttgcctgaa
     6121 gaaggtaagg ccattgaaga tgcagttaaa aaggttttgg atgcaggtat cagaactggt
     6181 gatttaggtg gttccaacag taccaccgaa gtcggtgatg ctgtcgccga agaagttaag
     6241 aaaatccttg cttaaaaaga ttctcttttt ttatgatatt tgtacataaa ctttataaat
     6301 gaaattcata atagaaacga cacgaaatta caaaatggaa tatgttcata gggtagacga
     6361 aactatatac gcaatctaca tacatttatc aagaaggaga aaaaggagga tgtaaaggaa
     6421 tacaggtaag caaattgata ctaatggctc aacgtgataa ggaaaaagaa ttgcacttta
     6481 acattaatat tgacaaggag gagggcacca cacaaaaagt taggtgtaac agaaaatcat
     6541 gaaactatga ttcctaattt atatattgga ggattttctc taaaaaaaaa aaatacaaca
     6601 aataaaaaac actcaatgac ctgaccattt gatggagttt aagtcaatac cttcttgaac
     6661 catttcccat aatggtgaaa gttccctcaa gaattttact ctgtcagaaa cggccttaac
     6721 gacgtagtcg atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     6781 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     6841 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     6901 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     6961 catgataata atggtttctt agacggatcg cttgcctgta acttacacgc gcctcgtatc
     7021 ttttaatgat ggaataattt gggaatttac tctgtgttta tttattttta tgttttgtat
     7081 ttggatttta gaaagtaaat aaagaaggta gaagagttac ggaatgaaga aaaaaaaata
     7141 aacaaaggtt taaaaaattt caacaaaaag cgtactttac atatatattt attagacaag
     7201 aaaagcagat taaatagata tacattcgat taacgataag taaaatgtaa aatcacagga
     7261 ttttcgtgtg tggtcttcta cacagacaag atgaaacaat tcggcattaa tacctgagag
     7321 caggaagagc aagataaaag gtagtatttg ttggcgatcc ccctagagtc ttttacatct
     7381 tcggaaaaca aaaactattt tttctttaat ttcttttttt actttctatt tttaatttat
     7441 atatttatat taaaaaattt aaattataat tatttttata gcacgtgatg aaaaggaccc
     7501 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     7561 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     7621 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     7681 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     7741 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     7801 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     7861 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     7921 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     7981 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     8041 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     8101 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     8161 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     8221 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     8281 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     8341 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     8401 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     8461 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     8521 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     8581 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     8641 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     8701 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     8761 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     8821 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctct
//
