LOCUS       sequence-381563-        6847 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cu2+-dependent expression of red fluorescent protein in yeast cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6847)
  AUTHORS   Utsumi R, Murata Y, Ito-Harashima S, Akai M, Miura N, Kuroda K, Ueda
            M, Kataoka M
  TITLE     Foci-forming regions of pyruvate kinase and enolase at the molecular
            surface incorporate proteins into yeast cytoplasmic metabolic 
            enzymes transiently assembling (META) bodies.
  JOURNAL   PLoS One. 2023 Apr 13;18(4):e0283002. doi: 
            10.1371/journal.pone.0283002. eCollection 2023.
  PUBMED    37053166
REFERENCE   2  (bases 1 to 6847)
  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: "PLoS One.";
            date: "2023-04-13"; volume: "18(4):e0283002. doi"; pages: " 
            10.1371/journal.pone.0283002. eCollection 2023"
COMMENT     Sequence Label: p426-Cup1p-FusionRed
FEATURES             Location/Qualifiers
     source          1..6847
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(22..40)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      81..1423
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    413..460
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        1450..1554
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1555..1623,1624..2415)
                     /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(1773..1792)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2586..3174
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3075..3094
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3328..3345
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3462..3483
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3498..3503,3504..3521,3522..3528)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3536..3552
                     /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     3541..3563
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3560..3576
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3560..3576
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3595..3615
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3597..3615
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3639..3873
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
     primer_bind     3888..3904
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3888..3904
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             3905..4603
                     /codon_start=1
                     /product="monomeric red fluorescent protein optimized for 
                     use as a fusion tag (Shemiakina et al., 2012)"
                     /label=FusionRed
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMPMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     VVEGGPLPFAFDILATSFMYGSRTFIKHPPGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKVRGVNFPANGPVMQKKTLGWEASTETMYPADGGLEGACDMAL
                     KLVGGGHLICNLETTYRSKKPATNLKMPGVYNVDHRLERIKEADDETYVEQHEVAVARY
                     STGGAGDGGK"
     terminator      4610..4857
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4626..4644)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4875..4894)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4876..4894)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4904..4921)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4904..4920)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4913..4935)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5061..5516
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5148..5167)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5358..5379
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5647..6450)
                     /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"
     promoter        complement(6451..6666)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(6690..6709)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6809..6831
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
       61 taataatggt ttcttagtat gatccaatat caaaggaaat gatagcattg aaggatgaga
      121 ctaatccaat tgaggagtgg cagcatatag aacagctaaa gggtagtgct gaaggaagca
      181 tacgataccc cgcatggaat gggataatat cacaggaggt actagactac ctttcatcct
      241 acataaatag acgcatataa gtacgcattt aagcataaac acgcactatg ccgttcttct
      301 catgtatata tatatacagg caacacgcag atataggtgc gacgtgaaca gtgagctgta
      361 tgtgcgcagc tcgcgttgca ttttcggaag cgctcgtttt cggaaacgct ttgaagttcc
      421 tattccgaag ttcctattct ctagaaagta taggaacttc agagcgcttt tgaaaaccaa
      481 aagcgctctg aagacgcact ttcaaaaaac caaaaacgca ccggactgta acgagctact
      541 aaaatattgc gaataccgct tccacaaaca ttgctcaaaa gtatctcttt gctatatatc
      601 tctgtgctat atccctatat aacctaccca tccacctttc gctccttgaa cttgcatcta
      661 aactcgacct ctacattttt tatgtttatc tctagtatta ctctttagac aaaaaaattg
      721 tagtaagaac tattcataga gtgaatcgaa aacaatacga aaatgtaaac atttcctata
      781 cgtagtatat agagacaaaa tagaagaaac cgttcataat tttctgacca atgaagaatc
      841 atcaacgcta tcactttctg ttcacaaagt atgcgcaatc cacatcggta tagaatataa
      901 tcggggatgc ctttatcttg aaaaaatgca cccgcagctt cgctagtaat cagtaaacgc
      961 gggaagtgga gtcaggcttt ttttatggaa gagaaaatag acaccaaagt agccttcttc
     1021 taaccttaac ggacctacag tgcaaaaagt tatcaagaga ctgcattata gagcgcacaa
     1081 aggagaaaaa aagtaatcta agatgctttg ttagaaaaat agcgctctcg ggatgcattt
     1141 ttgtagaaca aaaaagaagt atagattctt tgttggtaaa atagcgctct cgcgttgcat
     1201 ttctgttctg taaaaatgca gctcagattc tttgtttgaa aaattagcgc tctcgcgttg
     1261 catttttgtt ttacaaaaat gaagcacaga ttcttcgttg gtaaaatagc gctttcgcgt
     1321 tgcatttctg ttctgtaaaa atgcagctca gattctttgt ttgaaaaatt agcgctctcg
     1381 cgttgcattt ttgttctaca aaatgaagca cagatgcttc gttcaggtgg cacttttcgg
     1441 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     1501 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     1561 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     1621 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     1681 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     1741 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     1801 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     1861 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     1921 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     1981 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     2041 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     2101 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     2161 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     2221 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     2281 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     2341 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     2401 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     2461 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     2521 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     2581 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     2641 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     2701 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     2761 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     2821 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     2881 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     2941 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     3001 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     3061 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     3121 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     3181 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     3241 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     3301 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     3361 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     3421 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact
     3481 cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg
     3541 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gcgcgcaatt
     3601 aaccctcact aaagggaaca aaagctggag ctctagtact agttagaaaa agacattttt
     3661 gctgtcagtc actgtcaaga gattcttttg ctggcatttc ttctagaagc aaaaagagcg
     3721 atgcgtcttt tccgctgaac cgttccagca aaaaagacta ccaacgcaat atggattgtc
     3781 agaatcatat aaaagagaag caaataactc cttgtcttgt atcaattgca ttataatatc
     3841 ttcttgttag tgcaatatca tatagaagtc atcaaaaaac cccggattct agaactagtg
     3901 gatcatggtg agcgagctga ttaaggagaa catgcccatg aagctgtaca tggagggcac
     3961 cgtgaacaac caccacttca agtgcacatc cgagggcgaa ggcaagccct acgagggcac
     4021 ccagaccatg agaatcaagg tcgtcgaggg cggccctctc cccttcgcct tcgacatcct
     4081 ggctaccagc ttcatgtacg gcagcagaac cttcatcaag caccctccgg gcatccccga
     4141 cttctttaag cagtccttcc ctgagggctt cacatgggag agagtcacca catacgaaga
     4201 cgggggcgtg ctgaccgcta cccaggacac cagcctccag gacggctgcc tcatctacaa
     4261 cgtcaaggtt agaggggtga acttcccagc caacggccct gtgatgcaga agaaaacact
     4321 cggctgggag gcctccaccg agacgatgta ccccgctgac ggcggcctgg aaggcgcatg
     4381 tgacatggcc ctgaagctcg tgggcggggg ccacctgatc tgcaaccttg agaccacata
     4441 cagatccaag aaacccgcta cgaacctcaa gatgcccggc gtctacaacg tggaccacag
     4501 actggaaaga atcaaggagg ccgacgatga gacctacgtc gagcagcacg aggtggctgt
     4561 ggccagatac tctactggtg gcgctggtga tggaggtaaa taactcgagt catgtaatta
     4621 gttatgtcac gcttacattc acgccctccc cccacatccg ctctaaccga aaaggaagga
     4681 gttagacaac ctgaagtcta ggtccctatt tattttttta tagttatgtt agtattaaga
     4741 acgttattta tatttcaaat ttttcttttt tttctgtaca gacgcgtgta cgcatgtaac
     4801 attatactga aaaccttgct tgagaaggtt ttgggacgct cgaaggcttt aatttgcggc
     4861 cggtacccaa ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac
     4921 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc
     4981 ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     5041 gcagcctgaa tggcgaatgg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     5101 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     5161 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     5221 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     5281 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     5341 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     5401 ggtctattct tttgatttat aaggtatttt gccgatttcg gcctattggt taaaaaatga
     5461 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttcctg
     5521 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatagg gtaataactg
     5581 atataattaa attgaagctc taatttgtga gtttagtata catgcattta cttataatac
     5641 agttttttag ttttgctggc cgcatcttct caaatatgct tcccagcctg cttttctgta
     5701 acgttcaccc tctaccttag catcccttcc ctttgcaaat agtcctcttc caacaataat
     5761 aatgtcagat cctgtagaga ccacatcatc cacggttcta tactgttgac ccaatgcgtc
     5821 tcccttgtca tctaaaccca caccgggtgt cataatcaac caatcgtaac cttcatctct
     5881 tccacccatg tctctttgag caataaagcc gataacaaaa tctttgtcgc tcttcgcaat
     5941 gtcaacagta cccttagtat attctccagt agatagggag cccttgcatg acaattctgc
     6001 taacatcaaa aggcctctag gttcctttgt tacttcttct gccgcctgct tcaaaccgct
     6061 aacaatacct gggcccacca caccgtgtgc attcgtaatg tctgcccatt ctgctattct
     6121 gtatacaccc gcagagtact gcaatttgac tgtattacca atgtcagcaa attttctgtc
     6181 ttcgaagagt aaaaaattgt acttggcgga taatgccttt agcggcttaa ctgtgccctc
     6241 catggaaaaa tcagtcaaga tatccacatg tgtttttagt aaacaaattt tgggacctaa
     6301 tgcttcaact aactccagta attccttggt ggtacgaaca tccaatgaag cacacaagtt
     6361 tgtttgcttt tcgtgcatga tattaaatag cttggcagca acaggactag gatgagtagc
     6421 agcacgttcc ttatatgtag ctttcgacat gatttatctt cgtttcctgc aggtttttgt
     6481 tctgtgcagt tgggttaaga atactgggca atttcatgtt tcttcaacac tacatatgcg
     6541 tatatatacc aatctaagtc tgtgctcctt ccttcgttct tccttctgtt cggagattac
     6601 cgaatcaaaa aaatttcaag gaaaccgaaa tcaaaaaaaa gaataaaaaa aaaatgatga
     6661 attgaaaagg tggtatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     6721 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     6781 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     6841 tcatcac
//
