LOCUS       sequence-350991-        6516 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of eCFP for auxotrophic selection in the absence of 
            histidine.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6516)
  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 6516)
  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: pH-eCFP
FEATURES             Location/Qualifiers
     source          1..6516
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     165..183
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(221..243)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     343..362
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        509..696
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             697..1359
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(1618..2073)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1755..1776)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1967..1986
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2199..2221
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2213..2230
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2214..2230
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2237..2256
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2237..2255
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2282..2948
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     2818..2841
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             2973..3689
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYIYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGIDELYK"
     terminator      3710..3897
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(3765..3789)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        complement(3935..3953)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3974..3990)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3974..3990)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3987..4009)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3998..4014
                     /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(4022..4028,4029..4046,4047..4052))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4067..4088
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4205..4222)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4376..4964)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4456..4475)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5135..5926,5927..5995))
                     /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     5758..5777
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5996..6100)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    join(6130..6516,1..117)
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
ORIGIN
        1 ttacatcttc ggaaaacaaa aactattttt tctttaattt ctttttttac tttctatttt
       61 taatttatat atttatatta aaaaatttaa attataatta tttttatagc acgtgatgaa
      121 aaggacccta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
      181 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
      241 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
      301 cgtcagcggg tgttggcgga cgtcggggct ggcttaacta tgcggcatca gagcagattg
      361 tactgagagt gcaccataaa ttcccgtttt aagagcttgg tgagcgctag gagtcactgc
      421 caggtatcgt ttgaacacgg cattagtcag ggaagtcata acacagtcct ttcccgcaat
      481 tttctttttc tattactctt ggcctcctct agtacactct atattttttt atgcctcggt
      541 aatgattttc attttttttt ttccacctag cggatgactc tttttttttc ttagcgattg
      601 gcattatcac ataatgaatt atacattata taaagtaatg tgatttcttc gaagaatata
      661 ctaaaaaatg agcaggcaag ataaacgaag gcaaagatga cagagcagaa agccctagta
      721 aagcgtatta caaatgaaac caagattcag attgcgatct ctttaaaggg tggtccccta
      781 gcgatagagc actcgatctt cccagaaaaa gaggcagaag cagtagcaga acaggccaca
      841 caatcgcaag tgattaacgt ccacacaggt atagggtttc tggaccatat gatacatgct
      901 ctggccaagc attccggctg gtcgctaatc gttgagtgca ttggtgactt acacatagac
      961 gaccatcaca ccactgaaga ctgcgggatt gctctcggtc aagcttttaa agaggcccta
     1021 ggggccgtgc gtggagtaaa aaggtttgga tcaggatttg cgcctttgga tgaggcactt
     1081 tccagagcgg tggtagatct ttcgaacagg ccgtacgcag ttgtcgaact tggtttgcaa
     1141 agggagaaag taggagatct ctcttgcgag atgatcccgc attttcttga aagctttgca
     1201 gaggctagca gaattaccct ccacgttgat tgtctgcgag gcaagaatga tcatcaccgt
     1261 agtgagagtg cgttcaaggc tcttgcggtt gccataagag aagccacctc gcccaatggt
     1321 accaacgatg ttccctccac caaaggtgtt cttatgtagt gacaccgatt atttaaagct
     1381 gcagcatacg atatatatac atgtgtatat atgtatacct atgaatgtca gtaagtatgt
     1441 atacgaacag tatgatactg aagatgacaa ggtaatgcat cattctatac gtgtcattct
     1501 gaacgaggcg cgctttcctt ttttcttttt gctttttctt tttttttctc ttgaactcga
     1561 gatcctatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggaaa
     1621 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     1681 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     1741 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     1801 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     1861 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     1921 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     1981 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     2041 ccgccgcgct taatgcgccg ctacagggcg cgtccattcg ccattcaggc tgcgcaactg
     2101 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     2161 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     2221 gacggccagt gaattgtaat acgactcact atagggcgaa ttggagctcc accgcggtgg
     2281 ctcattatca atactgccat ttcaaagaat acgtaaataa ttaatagtag tgattttcct
     2341 aactttattt agtcaaaaaa ttagcctttt aattctgctg taacccgtac atgcccaaaa
     2401 tagggggcgg gttacacaga atatataaca tcgtaggtgt ctgggtgaac agtttattcc
     2461 tggcatccac taaatataat ggagcccgct ttttaagctg gcatccagaa aaaaaaagaa
     2521 tcccagcacc aaaatattgt tttcttcacc aaccatcagt tcataggtcc attctcttag
     2581 cgcaactaca gagaacaggg gcacaaacag gcaaaaaacg ggcacaacct caatggagtg
     2641 atgcaacctg cctggagtaa atgatgacac aaggcaattg acccacgcat gtatctatct
     2701 cattttctta caccttctat taccttctgc tctctctgat ttggaaaaag ctgaaaaaaa
     2761 aggttgaaac cagttccctg aaattattcc cctacttgac taataagtat ataaagacgg
     2821 taggtattga ttgtaattct gtaaatctat ttcttaaact tcttaaattc tacttttata
     2881 gttagtcttt tttttagttt taaaacacca agaacttagt ttcgaataaa cacacataaa
     2941 caaacaaagg tgacggtgct ggtttaatta acatgtctaa aggtgaagaa ttattcactg
     3001 gtgttgtccc aattttggtt gaattagatg gtgatgttaa tggtcacaaa ttttctgtct
     3061 ccggtgaagg tgaaggtgat gctacttacg gtaaattgac cttaaaattt atttgtacta
     3121 ctggtaaatt gccagttcca tggccaacct tagtcactac tttaacttgg ggtgttcaat
     3181 gtttttctag atacccagat catatgaaac aacatgactt tttcaagtct gccatgccag
     3241 aaggttatgt tcaagaaaga actatttttt tcaaagatga cggtaactac aagaccagag
     3301 ctgaagtcaa gtttgaaggt gataccttag ttaatagaat cgaattaaaa ggtattgatt
     3361 ttaaagaaga tggtaacatt ttaggtcaca aattggaata catttataac tctcacaatg
     3421 tttacatcac tgctgacaaa caaaagaatg gtatcaaagc taacttcaaa attagacaca
     3481 acattgaaga tggttctgtt caattagctg accattatca acaaaatact ccaattggtg
     3541 atggtccagt cttgttacca gacaaccatt acttatccac tcaatctgcc ttatccaaag
     3601 atccaaacga aaagagagac cacatggtct tgttagaatt tgttactgct gctggtatta
     3661 cccatggtat tgatgaattg tacaaataag gcgcgccact tctaaataag cgaatttctt
     3721 atgatttatg atttttatta ttaaataagt tataaaaaaa ataagtgtat acaaatttta
     3781 aagtgactct taggttttaa aacgaaaatt cttattcttg agtaactctt tcctgtaggt
     3841 caggttgctt tctcaggtat agtatgaggt cgctcttatt gaccacacct ctaccggtcg
     3901 acctggaggg ggggcccggt accagctttt gttcccttta gtgagggtta atttcgagct
     3961 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     4021 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     4081 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4141 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     4201 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4261 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4321 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4381 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4441 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4501 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4561 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4621 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4681 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4741 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     4801 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     4861 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     4921 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     4981 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5041 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5101 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5161 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5221 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     5281 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5341 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5401 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     5461 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     5521 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     5581 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     5641 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     5701 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     5761 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     5821 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     5881 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     5941 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6001 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6061 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6121 cacctgacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa tgatggaata
     6181 atttgggaat ttactctgta tttatttatt tttatgtttt gtatttggat tttagaaagt
     6241 aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa
     6301 atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc agattaaata
     6361 gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg tgtgtggtct
     6421 tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa gagcaagata
     6481 aaaggtagta tttgttggcg atccccctag agtctt
//
