LOCUS       sequence-450957-        7767 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for cytoplasmic aggregates.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7767)
  TITLE     Budding yeast expression plasmid for induced production of 
            fluorescently tagged cytoplasmic protein aggregates
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7767)
  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            "Unpublished"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*-mNeonGreen
FEATURES             Location/Qualifiers
     source          1..7767
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             661..1371
                     /codon_start=1
                     /product="bright monomeric yellow-green fluorescent protein
                     derived from LanYFP (Shaner et al., 2013)"
                     /label=mNeonGreen
                     /translation="MVSKGEEDNMASLPATHELYIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFTDVMGMDELYK"
     regulatory      916..925
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1509..1527)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(1693..1711)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1732..1748)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1732..1748)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1745..1767)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1756..1772
                     /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(1780..1786,1787..1804,1805..1810))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1825..1846
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1963..1980)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2134..2722)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2214..2233)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2893..3684,3685..3753))
                     /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     3516..3535
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3754..3858)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    3895..4398
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     4439..4457
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4495..4517)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4617..4636
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4660..4875
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             4876..5679
                     /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(5810..6265)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5947..5968)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6159..6178
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6391..6413
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6405..6422
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6406..6422
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6429..6448
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6429..6447
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        6611..6845
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
ORIGIN
        1 tcggaggaat gctctgctat ggaagactct ttggaacgtt gtttgggctt gatcgagtcg
       61 tgctatgact cgcaatccgt ctggtcctgt gttccagcta ccatttattg taataacgcc
      121 caattggctc cttaccaacg taccggcaga aacgtttacg atatcaggaa ggattgtgaa
      181 ggtggcaatt tgtgctaccc aacgttacaa gatatcgacg actacttaaa ccaggactac
      241 gtcaaagaag ctgtcggtgc ggaggttgac cactacgaat cctgtaactt cgatatcaac
      301 agaaatttcc tgtttgcggg tgattggatg aagccttacc acaccgccgt aacagatctt
      361 ttgaatcaag acctacccat tctggtatat gcaggcgata aagatttcat ctgtaactgg
      421 ttgggtaata aggcgtggac ggatgtcttg ccatggaagt acgacgaaga atttgcaagc
      481 caaaaagtac gtaactggac tgcttctatc accgacgagg tcgctggtga agtcaaatcc
      541 tacaagcact tcacctattt gagagtcttc aatggtggcc acatggttcc atttgacgtc
      601 cctgaaaacg ccttaagtat ggttaacgaa tggatccacg gtggtttctc cttactcgag
      661 atggtgagca agggcgagga ggataacatg gcctctctcc cagcgacaca tgagttatac
      721 atctttggct ccatcaacgg tgtggacttt gacatggtgg gtcagggcac cggcaatcca
      781 aatgatggtt atgaggagtt aaacctgaag tccaccaagg gtgacctcca gttctccccc
      841 tggattctgg tccctcatat cgggtatggc ttccatcagt acctgcccta ccctgacggg
      901 atgtcgcctt tccaggccgc catggtagat ggctccggct accaagtcca tcgcacaatg
      961 cagtttgaag atggtgcctc ccttactgtt aactaccgct acacctacga gggaagccac
     1021 atcaaaggag aggcccaggt gaaggggact ggtttccctg ctgacggtcc tgtgatgacc
     1081 aactcgctga ccgctgcgga ctggtgcagg tcgaagaaga cttaccccaa cgacaaaacc
     1141 atcatcagta cctttaagtg gagttacacc actggaaatg gcaagcgcta ccggagcact
     1201 gcgcggacca cctacacctt tgccaagcca atggcggcta actatctgaa gaaccagccg
     1261 atgtacgtgt tccgtaagac ggagctcaag cactccaaga ccgagctcaa cttcaaggag
     1321 tggcaaaagg cctttaccga tgtgatgggc atggacgagc tgtacaagta gaagctttgc
     1381 catagaatat ctggatccgg taccgggttg aagaaggaaa aagacagaaa cgacttaatt
     1441 acctacttga aaaaagcctg tgagtaaaca ggcccctttt cctttgtcga tatcatgtaa
     1501 ttagttatgt cacgcttaca ttcacgccct ccccccacat ccgctctaac cgaaaaggaa
     1561 ggagttagac aacctgaagt ctaggtccct atttattttt ttatagttat gttagtatta
     1621 agaacgttat ttatatttca aatttttctt ttttttctgt acaaacgcgt gtacggtacc
     1681 cagcttttgt tccctttagt gagggttaat tccgagcttg gcgtaatcat ggtcatagct
     1741 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     1801 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     1861 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     1921 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     1981 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     2041 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     2101 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     2161 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     2221 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2281 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     2341 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2401 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     2461 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     2521 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     2581 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     2641 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     2701 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     2761 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     2821 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     2881 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     2941 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     3001 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     3061 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     3121 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     3181 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     3241 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     3301 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     3361 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     3421 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     3481 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     3541 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     3601 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     3661 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     3721 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     3781 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     3841 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgggtcct tttcatcacg
     3901 tgctataaaa ataattataa tttaaatttt ttaatataaa tatataaatt aaaaatagaa
     3961 agtaaaaaaa gaaattaaag aaaaaatagt ttttgttttc cgaagatgta aaagactcta
     4021 gggggatcgc caacaaatac taccttttat cttgctcttc ctgctctcag gtattaatgc
     4081 cgaattgttt catcttgtct gtgtagaaga ccacacacga aaatcctgtg attttacatt
     4141 ttacttatcg ttaatcgaat gtatatctat ttaatctgct tttcttgtct aataaatata
     4201 tatgtaaagt acgctttttg ttgaaatttt ttaaaccttt gtttattttt ttttcttcat
     4261 tccgtaactc ttctaccttc tttatttact ttctaaaatc caaatacaaa acataaaaat
     4321 aaataaacac agagtaaatt cccaaattat tccatcatta aaagatacga ggcgcgtgta
     4381 agttacaggc aagcgatccg tctaagaaac cattattatc atgacattaa cctataaaaa
     4441 taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg
     4501 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     4561 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc
     4621 atcagagcag attgtactga gagtgcacca taccaccttt tcaattcatc attttttttt
     4681 tattcttttt tttgatttcg gtttccttga aatttttttg attcggtaat ctccgaacag
     4741 aaggaagaac gaaggaagga gcacagactt agattggtat atatacgcat atgtagtgtt
     4801 gaagaaacat gaaattgccc agtattctta acccaactgc acagaacaaa aacctgcagg
     4861 aaacgaagat aaatcatgtc gaaagctaca tataaggaac gtgctgctac tcatcctagt
     4921 cctgttgctg ccaagctatt taatatcatg cacgaaaagc aaacaaactt gtgtgcttca
     4981 ttggatgttc gtaccaccaa ggaattactg gagttagttg aagcattagg tcccaaaatt
     5041 tgtttactaa aaacacatgt ggatatcttg actgattttt ccatggaggg cacagttaag
     5101 ccgctaaagg cattatccgc caagtacaat tttttactct tcgaagacag aaaatttgct
     5161 gacattggta atacagtcaa attgcagtac tctgcgggtg tatacagaat agcagaatgg
     5221 gcagacatta cgaatgcaca cggtgtggtg ggcccaggta ttgttagcgg tttgaagcag
     5281 gcggcagaag aagtaacaaa ggaacctaga ggccttttga tgttagcaga attgtcatgc
     5341 aagggctccc tatctactgg agaatatact aagggtactg ttgacattgc gaagagcgac
     5401 aaagattttg ttatcggctt tattgctcaa agagacatgg gtggaagaga tgaaggttac
     5461 gattggttga ttatgacacc cggtgtgggt ttagatgaca agggagacgc attgggtcaa
     5521 cagtatagaa ccgtggatga tgtggtctct acaggatctg acattattat tgttggaaga
     5581 ggactatttg caaagggaag ggatgctaag gtagagggtg aacgttacag aaaagcaggc
     5641 tgggaagcat atttgagaag atgcggccag caaaactaaa aaactgtatt ataagtaaat
     5701 gcatgtatac taaactcaca aattagagct tcaatttaat tatatcagtt attaccctat
     5761 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcagga aattgtaagc
     5821 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     5881 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     5941 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     6001 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     6061 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     6121 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     6181 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     6241 cttaatgcgc cgctacaggg cgcgtccatt cgccattcag gctgcgcaac tgttgggaag
     6301 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     6361 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     6421 gtgaattgta atacgactca ctatagggcg aattggagct ccaccgcggt ggcggccgca
     6481 tgttcatgta tgtatctgta tttaaaacac ttttgtatta tttttcctca tatatgtgta
     6541 taggtttata cggatgattt aattattact tcaccaccct ttatttcagg ctgatatctt
     6601 agccttgtta ctagttagaa aaagacattt ttgctgtcag tcactgtcaa gagattcttt
     6661 tgctggcatt tcttctagaa gcaaaaagag cgatgcgtct tttccgctga accgttccag
     6721 caaaaaagac taccaacgca atatggattg tcagaatcat ataaaagaga agcaaataac
     6781 tccttgtctt gtgtcaattg cattataata tcttcttgtt agtgcaatat catatagaag
     6841 tcatcgaaat agatattaag aaaaacaaac tgtaacgaat tcatgatctc attgcaaaga
     6901 ccgttgggtc tagataagga cgttttgctg caagctgcgg aaaaatttgg tttggacctc
     6961 gacctggatc atctcttgaa ggagttggac tccaatgtat tggacgcttg ggcccaaata
     7021 gagcatttgt acccaaacca ggttatgagc cttgaaactt ccactaagcc aaaattccct
     7081 gaagcaatca aaacgaagaa agactgggac tttgtggtca agaatgacgc aattgaaaac
     7141 tatcagcttc gtgtcaacaa gattaaggac cctaaaatcc tgggcattga cccaaatgtc
     7201 acacagtaca cgggttactt ggatgtggaa gacgaggaca agcatttctt cttttggact
     7261 tttgaaagta gaaacgatcc tgcaaaggat ccggtcatcc tttggttgaa cgggggtcca
     7321 ggttgttctt cactaaccgg gctgttcttt gaattaggac cctcatccat tggacctgat
     7381 ttgaaaccca tcgggaaccc ttactcttgg aacagcaatg ccaccgtgat cttccttgac
     7441 cagcctgtca acgttgggtt ctcgtattcc gggtcctcag gtgtttccaa cactgtcgcc
     7501 gctggtaagg atgtctataa cttcttggag ttgttcttcg atcagttccc tgaatacgtc
     7561 aacaagggcc aagatttcca catcgctagg gaatcctacg ccggccatta catccctgtt
     7621 tttgcctctg aaattttgtc tcacaaggac agaaacttca acttaacctc cgtcttgatc
     7681 ggaaatggcc tcactgaccc attgactcag tataactatt acgaaccaat ggcctgtggt
     7741 gaaggtggcg aaccatctgt tttgccc
//
