LOCUS       sequence-107475-        8284 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  harbours marker genes HIS3, LEU2 and URA3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8284)
  AUTHORS   Mulleder M, Campbell K, Matsarskaia O, Eckerstorfer F, Ralser M
  TITLE     Saccharomyces cerevisiae single-copy plasmids for auxotrophy 
            compensation, multiple marker selection, and for designing 
            metabolically cooperating communities.
  JOURNAL   F1000Res. 2016 Sep 20;5:2351. eCollection 2016.
  PUBMED    27830062
REFERENCE   2  (bases 1 to 8284)
  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: "F1000Res.";
            date: "2016-09-20"; volume: "5"; pages: "2351. eCollection 2016"
COMMENT     Sequence Label: pHLU
FEATURES             Location/Qualifiers
     source          1..8284
                     /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        507..694
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             695..1357
                     /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"
     CDS             1716..2519
                     /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        2897..3304
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             3305..4399
                     /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"
     rep_origin      complement(4934..5393)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5071..5092)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5283..5302
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5519..5541
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5533..5550
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5534..5550
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5557..5576
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5557..5575
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    5584..5691
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     promoter        complement(5703..5721)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5742..5758)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5742..5758)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5755..5777)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5766..5782
                     /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(5790..5796,5797..5814,5815..5820))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5835..5856
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5973..5990)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6144..6732)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6224..6243)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6903..7694,7695..7763))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7526..7545
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7764..7868)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    join(7898..8284,1..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 gcaccataat tccgttttaa gagcttggtg agcgctagga gtcactgcca
      421 ggtatcgttt gaacacggca ttagtcaggg aagtcataac acagtccttt cccgcaattt
      481 tctttttcta ttactcttgg cctcctctag tacactctat atttttttat gcctcggtaa
      541 tgattttcat tttttttttt ccacctagcg gatgactctt tttttttctt agcgattggc
      601 attatcacat aatgaattat acattatata aagtaatgtg atttcttcga agaatatact
      661 aaaaaatgag caggcaagat aaacgaaggc aaagatgaca gagcagaaag ccctagtaaa
      721 gcgtattaca aatgaaacca agattcagat tgcgatctct ttaaagggtg gtcccctagc
      781 gatagagcac tcgatcttcc cagaaaaaga ggcagaagca gtagcagaac aggccacaca
      841 atcgcaagtg attaacgtcc acacaggtat agggtttctg gaccatatga tacatgctct
      901 ggccaagcat tccggctggt cgctaatcgt tgagtgcatt ggtgacttac acatagacga
      961 ccatcacacc actgaagact gcgggattgc tctcggtcaa gcttttaaag aggccctagg
     1021 ggccgtgcgt ggagtaaaaa ggtttggatc aggatttgcg cctttggatg aggcactttc
     1081 cagagcggtg gtagatcttt cgaacaggcc gtacgcagtt gtcgaacttg gtttgcaaag
     1141 ggagaaagta ggagatctct cttgcgagat gatcccgcat tttcttgaaa gctttgcaga
     1201 ggctagcaga attaccctcc acgttgattg tctgcgaggc aagaatgatc atcaccgtag
     1261 tgagagtgcg ttcaaggctc ttgcggttgc cataagagaa gccacctcgc ccaatggtac
     1321 caacgatgtt ccctccacca aaggtgttct tatgtagtga caccgattat ttaaagctgc
     1381 agcatacgat atatatacat gtgtatatat gtatacctat gaatgtcagt aagtatgtat
     1441 acgaacagta tgatactgaa gatgacaagg taatgcatca ttctatacgt gtcattctga
     1501 acgaggcgcg ctttcctttt ttctttttgc tttttctttt tttttctctt gaactcgacg
     1561 attcggtaat ctccgaacag aaggaagaac gaaggaagga gcacagactt agattggtat
     1621 atatacgcat atgtagtgtt gaagaaacat gaaattgccc agtattctta acccaactgc
     1681 acagaacaaa aacctgcagg aaacgaagat aaatcatgtc gaaagctaca tataaggaac
     1741 gtgctgctac tcatcctagt cctgttgctg ccaagctatt taatatcatg cacgaaaagc
     1801 aaacaaactt gtgtgcttca ttggatgttc gtaccaccaa ggaattactg gagttagttg
     1861 aagcattagg tcccaaaatt tgtttactaa aaacacatgt ggatatcttg actgattttt
     1921 ccatggaggg cacagttaag ccgctaaagg cattatccgc caagtacaat tttttactct
     1981 tcgaagacag aaaatttgct gacattggta atacagtcaa attgcagtac tctgcgggtg
     2041 tatacagaat agcagaatgg gcagacatta cgaatgcaca cggtgtggtg ggcccaggta
     2101 ttgttagcgg tttgaagcag gcggcagaag aagtaacaaa ggaacctaga ggccttttga
     2161 tgttagcaga attgtcatgc aagggctccc tatctactgg agaatatact aagggtactg
     2221 ttgacattgc gaagagcgac aaagattttg ttatcggctt tattgctcaa agagacatgg
     2281 gtggaagaga tgaaggttac gattggttga ttatgacacc cggtgtgggt ttagatgaca
     2341 agggagacgc attgggtcaa cagtatagaa ccgtggatga tgtggtctct acaggatctg
     2401 acattattat tgttggaaga ggactatttg caaagggaag ggatgctaag gtagagggtg
     2461 aacgttacag aaaagcaggc tgggaagcat atttgagaag atgcggccag caaaactaaa
     2521 aaactgtatt ataagtaaat gcatgtatac taaactcaca aattagagct tcaatttaat
     2581 tatatcagtt attaccctat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     2641 cgcatcaggc gcgccatcga ggagaacttc tagtatatct acatacctaa tattattgcc
     2701 ttattaaaaa tggaatccca acaattacat caaaatccac attctcttca aaatcaattg
     2761 tcctgtactt ccttgttcat gtgtgttcaa aaacgttata tttataggat aattatactc
     2821 tatttctcaa caagtaattg gttgtttggc cgagcggtct aaggcgcctg attcaagaaa
     2881 tatcttgacc gcagttaact gtgggaatac tcaggtatcg taagatgcaa gagttcgaat
     2941 ctcttagcaa ccattatttt tttcctcaac ataacgagaa cacacagggg cgctatcgca
     3001 cagaatcaaa ttcgatgact ggaaattttt tgttaatttc agaggtcgcc tgacgcatat
     3061 acctttttca actgaaaaat tgggagaaaa aggaaaggtg agagcgccgg aaccggcttt
     3121 tcatatagaa tagagaagcg ttcatgacta aatgcttgca tcacaatact tgaagttgac
     3181 aatattattt aaggacctat tgttttttcc aataggtggt tagcaatcgt cttactttct
     3241 aacttttctt accttttaca tttcagcaat atatatatat atatttcaag gatataccat
     3301 tctaatgtct gcccctaaga agatcgtcgt tttgccaggt gaccacgttg gtcaagaaat
     3361 cacagccgaa gccattaagg ttcttaaagc tatttctgat gttcgttcca atgtcaagtt
     3421 cgatttcgaa aatcatttaa ttggtggtgc tgctatcgat gctacaggtg ttccacttcc
     3481 agatgaggcg ctggaagcct ccaagaaggc tgatgccgtt ttgttaggtg ctgtgggtgg
     3541 tcctaaatgg ggtaccggta gtgttagacc tgaacaaggt ttactaaaaa tccgtaaaga
     3601 acttcaattg tacgccaact taagaccatg taactttgca tccgactctc ttttagactt
     3661 atctccaatc aagccacaat ttgctaaagg tactgacttc gttgttgtca gagaattagt
     3721 gggaggtatt tactttggta agagaaagga agacgatggt gatggtgtcg cttgggatag
     3781 tgaacaatac accgttccag aagtgcaaag aatcacaaga atggccgctt tcatggccct
     3841 acaacatgag ccaccattgc ctatttggtc cttggataaa gctaatgttt tggcctcttc
     3901 aagattatgg agaaaaactg tggaggaaac catcaagaac gaattcccta cattgaaggt
     3961 tcaacatcaa ttgattgatt ctgccgccat gatcctagtt aagaacccaa cccacctaaa
     4021 tggtattata atcaccagca acatgtttgg tgatatcatc tccgatgaag cctccgttat
     4081 cccaggttcc ttgggtttgt tgccatctgc gtccttggcc tctttgccag acaagaacac
     4141 cgcatttggt ttgtacgaac catgccacgg ttctgctcca gatttgccaa agaataaggt
     4201 caaccctatc gccactatct tgtctgctgc aatgatgttg aaattgtcat tgaacttgcc
     4261 tgaagaaggt aaggccattg aagatgcagt taaaaaggtt ttggatgcag gtatcagaac
     4321 tggtgattta ggtggttcca acagtaccac cgaagtcggt gatgctgtcg ccgaagaagt
     4381 taagaaaatc cttgcttaaa aagattctct ttttttatga tatttgtaca taaactttat
     4441 aaatgaaatt cataatagaa acgacacgaa attacaaaat ggaatatgtt catagggtag
     4501 acgaaactat atacgcaatc tacatacatt tatcaagaag gagaaaaagg aggatgtaaa
     4561 ggaatacagg taagcaaatt gatactaatg gctcaacgtg ataaggaaaa agaattgcac
     4621 tttaacatta atattgacaa ggaggagggc accacacaaa aagttaggtg taacagaaaa
     4681 tcatgaaact atgattccta atttatatat tggaggattt tctctaaaaa aaaaaaaata
     4741 caacaaataa aaaacactca atgacctgac catttgatgg agtttaagtc aataccttct
     4801 tgaaccattt cccataatgg tgaaagttcc ctcaagaatt ttactctgtc agaaacggcc
     4861 ttacgacgta gtcgaggatc ctatgcggtg tgaaataccg cacagatgcg taaggagaaa
     4921 ataccgcatc aggaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt
     4981 taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa
     5041 gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag
     5101 aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt
     5161 gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac
     5221 cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag
     5281 gaagggaaga aagcgaaagg agcgggcgct agggcgctgg ctggcaagtg tagcggtcac
     5341 gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtccattcg
     5401 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
     5461 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc
     5521 cagtcacgac gttgtaaaac gacggccagt gaattgtaat acgactcact atagggcgaa
     5581 ttggagctcc accgcggtgg cggccgctct agaactagtg ggtcccccgg gctgcaggaa
     5641 ttcgatatca agcttatcga taccgtcgac ctggaggggg ggcccggtac cagcttttgt
     5701 tccctttagt gagggttaat ttcgagcttg gcgtaatcat ggtcatagct gtttcctgtg
     5761 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     5821 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     5881 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5941 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     6001 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     6061 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     6121 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     6181 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     6241 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     6301 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     6361 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6421 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6481 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6541 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     6601 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6661 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6721 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6781 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6841 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6901 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6961 atagttgcct gactgcccgt cgtgtagata actacgatac gggagggctt accatctggc
     7021 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     7081 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     7141 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     7201 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     7261 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgaaaaaaa
     7321 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7381 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7441 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7501 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7561 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7621 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7681 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7741 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     7801 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7861 gttccgcgca catttccccg aaaagtgcca cctgacggat cgcttgcctg taacttacac
     7921 gcgcctcgta tcttttaatg atggaataat ttgggaattt actctgtatt tatttatttt
     7981 tatgttttgt atttggattt tagaaagtaa ataaagaagg tagaagagtt acggaatgaa
     8041 gaaaaaaaaa taaacaaagg tttaaaaaat ttcaacaaaa agcgtacttt acatatatat
     8101 ttattagaca agaaaagcag attaaataga tatacattcg attaacgata agtaaaatgt
     8161 aaaatcacag gattttcgtg tgtggtcttc tacacagaca agatgaaaca attcggcatt
     8221 aatacctgag agcaggaaga gcaagataaa aggtagtatt tgttggcgat ccccctagag
     8281 tctt
//
