LOCUS       sequence-328574-        7678 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mRuby3 expression under control of the GPD promoter in yeast.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7678)
  AUTHORS   Harel I, Chen YR, Ziv I, Singh PP, Heinzer D, Navarro Negredo P, 
            Goshtchevsky U, Wang W, Astre G, Moses E, McKay A, Machado BE, 
            Hebestreit K, Yin S, Sanchez Alvarado A, Jarosz DF, Brunet A
  TITLE     Identification of protein aggregates in the aging vertebrate brain 
            with prion-like and phase-separation properties.
  JOURNAL   Cell Rep. 2024 Jun 25;43(6):112787. doi: 
            10.1016/j.celrep.2023.112787. Epub 2024 May 28.
  PUBMED    38810650
REFERENCE   2  (bases 1 to 7678)
  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: "Cell Rep.";
            date: "2024-06-25"; volume: "43(6):112787. doi"; pages: " 
            10.1016/j.celrep.2023.112787. Epub 2024 May 28"
COMMENT     Sequence Label: pAG415GPD-mRuby3
FEATURES             Location/Qualifiers
     source          1..7678
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    6..509
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        546..650
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(651..719,720..1511)
                     /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(869..888)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1682..2270
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2171..2190
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2424..2441
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2558..2579
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2594..2599,2600..2617,2618..2624)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2632..2648
                     /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     2637..2659
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2656..2672
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2656..2672
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2691..2711
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2693..2711
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2736..3379
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     3273..3296
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     3385..3401
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3385..3401
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    3409..3433
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3451..4161
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
     protein_bind    complement(4166..4190)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(4223..4239)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4224..4240)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      4242..4489
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4258..4276)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4507..4526)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4508..4526)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4536..4553)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4536..4552)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4545..4567)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4693..5148
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4780..4799)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4990..5011
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5449..5856
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             5857..6951
                     /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(7445..7464)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7564..7586
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7624..7642)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ggacggatcg cttgcctgta acttacacgc gcctcgtatc ttttaatgat ggaataattt
       61 gggaatttac tctgtgttta tttattttta tgttttgtat ttggatttta gaaagtaaat
      121 aaagaaggta gaagagttac ggaatgaaga aaaaaaaata aacaaaggtt taaaaaattt
      181 caacaaaaag cgtactttac atatatattt attagacaag aaaagcagat taaatagata
      241 tacattcgat taacgataag taaaatgtaa aatcacagga ttttcgtgtg tggtcttcta
      301 cacagacaag atgaaacaat tcggcattaa tacctgagag caggaagagc aagataaaag
      361 gtagtatttg ttggcgatcc ccctagagtc ttttacatct tcggaaaaca aaaactattt
      421 tttctttaat ttcttttttt actttctatt tttaatttat atatttatat taaaaaattt
      481 aaattataat tatttttata gcacgtgatg aaaaggaccc aggtggcact tttcggggaa
      541 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
      601 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
      661 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
      721 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
      781 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
      841 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
      901 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
      961 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     1021 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     1081 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     1141 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     1201 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     1261 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     1321 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     1381 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     1441 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     1501 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     1561 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     1621 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     1681 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     1741 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     1801 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     1861 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     1921 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     1981 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     2041 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     2101 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     2161 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     2221 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     2281 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     2341 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     2401 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa
     2461 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     2521 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt
     2581 aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg
     2641 gataacaatt tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc
     2701 ctcactaaag ggaacaaaag ctggagctca gtttatcatt atcaatactc gccatttcaa
     2761 agaatacgta aataattaat agtagtgatt ttcctaactt tatttagtca aaaaattagc
     2821 cttttaattc tgctgtaacc cgtacatgcc caaaataggg ggcgggttac acagaatata
     2881 taacatcgta ggtgtctggg tgaacagttt attcctggca tccactaaat ataatggagc
     2941 ccgcttttta agctggcatc cagaaaaaaa aagaatccca gcaccaaaat attgttttct
     3001 tcaccaacca tcagttcata ggtccattct cttagcgcaa ctacagagaa caggggcaca
     3061 aacaggcaaa aaacgggcac aacctcaatg gagtgatgca acctgcctgg agtaaatgat
     3121 gacacaaggc aattgaccca cgcatgtatc tatctcattt tcttacacct tctattacct
     3181 tctgctctct ctgatttgga aaaagctgaa aaaaaaggtt gaaaccagtt ccctgaaatt
     3241 attcccctac ttgactaata agtatataaa gacggtaggt attgattgta attctgtaaa
     3301 tctatttctt aaacttctta aattctactt ttatagttag tctttttttt agttttaaaa
     3361 caccagaact tagtttcgac ggattctaga actagtggat cccccatcac aagtttgtac
     3421 aaaaaagcag gcttcgaagg agataacaaa atggtgtcta agggcgaaga gctgatcaag
     3481 gaaaatatgc gtatgaaggt ggtcatggaa ggttcggtca acggccacca attcaaatgc
     3541 acaggtgaag gagaaggcag accgtacgag ggagtgcaaa ccatgaggat caaagtcatc
     3601 gagggaggac ccctgccatt tgcctttgac attcttgcca cgtcgttcat gtatggcagc
     3661 cgtaccttta tcaagtaccc ggccgacatc cctgatttct ttaaacagtc ctttcctgag
     3721 ggttttactt gggaaagagt tacgagatac gaagatggtg gagtcgtcac cgtcacgcag
     3781 gacaccagcc ttgaggatgg cgagctcgtc tacaacgtca aggtcagagg ggtaaacttt
     3841 ccctccaatg gtcccgtgat gcagaagaag accaagggtt gggagcctaa tacagagatg
     3901 atgtatccag cagatggtgg tctgagagga tacactgaca tcgcactgaa agttgatggt
     3961 ggtggccatc tgcactgcaa cttcgtgaca acttacaggt caaaaaagac cgtcgggaac
     4021 atcaagatgc ccggtgtcca tgccgttgat caccgcctgg aaaggatcga ggagagtgac
     4081 aatgaaacct acgtagtgca aagagaagtg gcagttgcca aatacagcaa ccttggtggt
     4141 ggcatggacg agctgtacaa gtaagaccca gctttcttgt acaaagtggt gatgggctgc
     4201 aggaattcga tatcaagctt atcgataccg tcgacctcga gtcatgtaat tagttatgtc
     4261 acgcttacat tcacgccctc cccccacatc cgctctaacc gaaaaggaag gagttagaca
     4321 acctgaagtc taggtcccta tttatttttt tatagttatg ttagtattaa gaacgttatt
     4381 tatatttcaa atttttcttt tttttctgta cagacgcgtg tacgcatgta acattatact
     4441 gaaaaccttg cttgagaagg ttttgggacg ctcgaaggct ttaatttgcg gccggtaccc
     4501 aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt
     4561 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     4621 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     4681 aatggcgaat ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     4741 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     4801 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     4861 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     4921 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     4981 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     5041 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     5101 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttcc tgatgcggta
     5161 ttttctcctt acgcatctgt gcggtatttc acaccgcata tcgaccggtc gaggagaact
     5221 tctagtatat ctacatacct aatattattg ccttattaaa aatggaatcc caacaattac
     5281 atcaaaatcc acattctctt caaaatcaat tgtcctgtac ttccttgttc atgtgtgttc
     5341 aaaaacgtta tatttatagg ataattatac tctatttctc aacaagtaat tggttgtttg
     5401 gccgagcggt ctaaggcgcc tgattcaaga aatatcttga ccgcagttaa ctgtgggaat
     5461 actcaggtat cgtaagatgc aagagttcga atctcttagc aaccattatt tttttcctca
     5521 acataacgag aacacacagg ggcgctatcg cacagaatca aattcgatga ctggaaattt
     5581 tttgttaatt tcagaggtcg cctgacgcat ataccttttt caactgaaaa attgggagaa
     5641 aaaggaaagg tgagagcgcc ggaaccggct tttcatatag aatagagaag cgttcatgac
     5701 taaatgcttg catcacaata cttgaagttg acaatattat ttaaggacct attgtttttt
     5761 ccaataggtg gttagcaatc gtcttacttt ctaacttttc ttacctttta catttcagca
     5821 atatatatat atatatttca aggatatacc attctaatgt ctgcccctaa gaagatcgtc
     5881 gttttgccag gtgaccacgt tggtcaagaa atcacagccg aagccattaa ggttcttaaa
     5941 gctatttctg atgttcgttc caatgtcaag ttcgatttcg aaaatcattt aattggtggt
     6001 gctgctatcg atgctacagg tgttccactt ccagatgagg cgctggaagc ctccaagaag
     6061 gctgatgccg ttttgttagg tgctgtgggt ggtcctaaat ggggtaccgg tagtgttaga
     6121 cctgaacaag gtttactaaa aatccgtaaa gaacttcaat tgtacgccaa cttaagacca
     6181 tgtaactttg catccgactc tcttttagac ttatctccaa tcaagccaca atttgctaaa
     6241 ggtactgact tcgttgttgt cagagaatta gtgggaggta tttactttgg taagagaaag
     6301 gaagacgatg gtgatggtgt cgcttgggat agtgaacaat acaccgttcc agaagtgcaa
     6361 agaatcacaa gaatggccgc tttcatggcc ctacaacatg agccaccatt gcctatttgg
     6421 tccttggata aagctaatgt tttggcctct tcaagattat ggagaaaaac tgtggaggaa
     6481 accatcaaga acgaattccc tacattgaag gttcaacatc aattgattga ttctgccgcc
     6541 atgatcctag ttaagaaccc aacccaccta aatggtatta taatcaccag caacatgttt
     6601 ggtgatatca tctccgatga agcctccgtt atcccaggtt ccttgggttt gttgccatct
     6661 gcgtccttgg cctctttgcc agacaagaac accgcatttg gtttgtacga accatgccac
     6721 ggttctgctc cagatttgcc aaagaataag gtcaacccta tcgccactat cttgtctgct
     6781 gcaatgatgt tgaaattgtc attgaacttg cctgaagaag gtaaggccat tgaagatgca
     6841 gttaaaaagg ttttggatgc aggtatcaga actggtgatt taggtggttc caacagtacc
     6901 accgaagtcg gtgatgctgt cgccgaagaa gttaagaaaa tccttgctta aaaagattct
     6961 ctttttttat gatatttgta cataaacttt ataaatgaaa ttcataatag aaacgacacg
     7021 aaattacaaa atggaatatg ttcatagggt agacgaaact atatacgcaa tctacataca
     7081 tttatcaaga aggagaaaaa ggaggatgta aaggaataca ggtaagcaaa ttgatactaa
     7141 tggctcaacg tgataaggaa aaagaattgc actttaacat taatattgac aaggaggagg
     7201 gcaccacaca aaaagttagg tgtaacagaa aatcatgaaa ctatgattcc taatttatat
     7261 attggaggat tttctctaaa aaaaaaaaaa tacaacaaat aaaaaacact caatgacctg
     7321 accatttgat ggagtttaag tcaatacctt cttgaaccat ttcccataat ggtgaaagtt
     7381 ccctcaagaa ttttactctg tcagaaacgg ccttaacgac gtagtcgata tggtgcactc
     7441 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
     7501 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     7561 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa
     7621 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttctta
//
