LOCUS       sequence-350277-        5707 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of mRuby2 for auxotrophic selection in the absence of 
            uracil.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5707)
  AUTHORS   Yu JSL, Correia-Melo C, Zorrilla F, Herrera-Dominguez L, Wu MY, 
            Hartl J, Campbell K, Blasche S, Kreidl M, Egger AS, Messner CB, 
            Demichev V, Freiwald A, Mulleder M, Howell M, Berman J, Patil KR, 
            Alam MT, Ralser M
  TITLE     Microbial communities form rich extracellular metabolomes that 
            foster metabolic interactions and promote drug tolerance.
  JOURNAL   Nat Microbiol. 2022 Apr;7(4):542-555. doi: 
            10.1038/s41564-022-01072-5. Epub 2022 Mar 21.
  PUBMED    35314781
REFERENCE   2  (bases 1 to 5707)
  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"; doi: 
            "10.1038/s41564-022-01072-5"; journalName: "Nat Microbiol"; date: 
            "2022-04"; volume: "7"; issue: "4"; pages: "542-555"
COMMENT     Sequence Label: pU-mRuby2
FEATURES             Location/Qualifiers
     source          1..5707
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     282..301
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        320..986
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     856..879
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             1022..1732
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     terminator      1756..1943
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1811..1835)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     misc_feature    2010..2513
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     CDS             2873..3676
                     /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"
     primer_bind     complement(3742..3761)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3743..3761)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3848..3865)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4019..4607)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4099..4118)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4778..5569,5570..5638))
                     /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     5401..5420
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5639..5707,1..36))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
       61 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
      121 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
      181 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
      241 gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt
      301 actgagagtg caccatatgt cattatcaat actgccattt caaagaatac gtaaataatt
      361 aatagtagtg attttcctaa ctttatttag tcaaaaaatt agccttttaa ttctgctgta
      421 acccgtacat gcccaaaata gggggcgggt tacacagaat atataacatc gtaggtgtct
      481 gggtgaacag tttattcctg gcatccacta aatataatgg agcccgcttt ttaagctggc
      541 atccagaaaa aaaaagaatc ccagcaccaa aatattgttt tcttcaccaa ccatcagttc
      601 ataggtccat tctcttagcg caactacaga gaacaggggc acaaacaggc aaaaaacggg
      661 cacaacctca atggagtgat gcaacctgcc tggagtaaat gatgacacaa ggcaattgac
      721 ccacgcatgt atctatctca ttttcttaca ccttctatta ccttctgctc tctctgattt
      781 ggaaaaagct gaaaaaaaag gttgaaacca gttccctgaa attattcccc tacttgacta
      841 ataagtatat aaagacggta ggtattgatt gtaattctgt aaatctattt cttaaacttc
      901 ttaaattcta cttttatagt tagtcttttt tttagtttta aaacaccaag aacttagttt
      961 cgaataaaca cacataaaca aacaaagtcg acggatcggt gacggtgctg gtttaattaa
     1021 catggtgtcc aaaggagagg agttaatcaa ggaaaacatg agaatgaaag ttgtcatgga
     1081 gggctccgtt aatggtcacc aattcaagtg tacaggggaa ggtgaaggta atccttacat
     1141 gggtacacaa actatgagaa ttaaagtaat tgaaggcgga ccactaccat ttgcatttga
     1201 cattctggca acgtcattca tgtacggatc acgaactttc atcaagtacc ctaaaggtat
     1261 accagacttt ttcaagcaat cttttccaga gggttttaca tgggaaaggg ttacaagata
     1321 cgaagatggg ggtgtcgtca cagttatgca agatacttca ttagaagatg gctgccttgt
     1381 ctatcatgtg caagtaagag gggtgaattt tccttctaac ggacctgtga tgcagaaaaa
     1441 gaccaaaggt tgggaaccaa atactgaaat gatgtaccca gctgatggag gtttgagagg
     1501 ctacacacac atggcgctta aagttgatgg tggaggtcat ttgtcttgta gttttgttac
     1561 cacttatcgt tctaaaaaga ctgttggcaa tatcaaaatg ccaggaatac atgctgtaga
     1621 ccacagacta gaaagactcg aagagagcga taacgaaatg ttcgttgtac agagagagca
     1681 tgccgtagcc aaatttgctg gcttaggcgg tggtatggat gaattgtata agtaaggcgc
     1741 gccacttcta aataagcgaa tttcttatga tttatgattt ttattattaa ataagttata
     1801 aaaaaaataa gtgtatacaa attttaaagt gactcttagg ttttaaaacg aaaattctta
     1861 ttcttgagta actctttcct gtaggtcagg ttgctttctc aggtatagta tgaggtcgct
     1921 cttattgacc acacctctac cggcagatcc gctagggata acagggtaat atagatctgt
     1981 ttagcttgcc tcgtccccgc cgggtcaccg atcgcttgcc tgtaacttac acgcgcctcg
     2041 catcttttaa tgatggaata atttgggaat ttactctgta tttatttatt tttatgtttt
     2101 gtatttggat tttagaaagt aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa
     2161 aataaacaaa ggtttaaaaa atttcaacaa aaagcgtact ttacatatat atttattaga
     2221 caagaaaagc agattaaata gatatacatt cgattaacga taagtaaaat gtaaaatcac
     2281 aggattttcg tgtgtggtct tctacacaga caagatgaaa caattcggca ttaatacctg
     2341 agagcaggaa gagcaagata aaaggtagta tttgttggcg atccccctag agtcttttac
     2401 atcttcggaa aacaaaaact attttttctt taatttcttt ttttactttc tatttttaat
     2461 ttatatattt atattaaaaa atttaaatta taattatttt tatagcacgt gatgaaaagg
     2521 accctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     2581 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     2641 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     2701 agcgggtgtt ggcggcgatt cggtaatctc cgaacagaag gaagaacgaa ggaaggagca
     2761 cagacttaga ttggtatata tacgcatatg tagtgttgaa gaaacatgaa attgcccagt
     2821 attcttaacc caactgcaca gaacaaaaac ctgcaggaaa cgaagataaa tcatgtcgaa
     2881 agctacatat aaggaacgtg ctgctactca tcctagtcct gttgctgcca agctatttaa
     2941 tatcatgcac gaaaagcaaa caaacttgtg tgcttcattg gatgttcgta ccaccaagga
     3001 attactggag ttagttgaag cattaggtcc caaaatttgt ttactaaaaa cacatgtgga
     3061 tatcttgact gatttttcca tggagggcac agttaagccg ctaaaggcat tatccgccaa
     3121 gtacaatttt ttactcttcg aagacagaaa atttgctgac attggtaata cagtcaaatt
     3181 gcagtactct gcgggtgtat acagaatagc agaatgggca gacattacga atgcacacgg
     3241 tgtggtgggc ccaggtattg ttagcggttt gaagcaggcg gcagaagaag taacaaagga
     3301 acctagaggc cttttgatgt tagcagaatt gtcatgcaag ggctccctat ctactggaga
     3361 atatactaag ggtactgttg acattgcgaa gagcgacaaa gattttgtta tcggctttat
     3421 tgctcaaaga gacatgggtg gaagagatga aggttacgat tggttgatta tgacacccgg
     3481 tgtgggttta gatgacaagg gagacgcatt gggtcaacag tatagaaccg tggatgatgt
     3541 ggtctctaca ggatctgaca ttattattgt tggaagagga ctatttgcaa agggaaggga
     3601 tgctaaggta gagggtgaac gttacagaaa agcaggctgg gaagcatatt tgagaagatg
     3661 cggccagcaa aactaagagc tcgaattcat cgatgatatc agatccacta gtggcctatg
     3721 cggccgcgga tctgccggtc tccctatagt gagtcgtatt aatttcgata agccaggtta
     3781 acctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     3841 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     3901 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     3961 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     4021 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     4081 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     4141 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     4201 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     4261 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     4321 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4381 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4441 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4501 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4561 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4621 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     4681 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     4741 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     4801 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     4861 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     4921 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     4981 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     5041 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     5101 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     5161 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     5221 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     5281 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     5341 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     5401 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     5461 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     5521 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     5581 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     5641 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     5701 tatttga
//
