LOCUS       sequence-107491-        5953 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  harbours marker gene LEU2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5953)
  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 5953)
  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: pL
FEATURES             Location/Qualifiers
     source          1..5953
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     164..182
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(220..242)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        566..973
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             974..2068
                     /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(2603..3062)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2740..2761)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2952..2971
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3188..3210
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3202..3219
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3203..3219
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3226..3245
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3226..3244
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    3253..3360
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     promoter        complement(3372..3390)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3411..3427)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3411..3427)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3424..3446)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3435..3451
                     /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(3459..3465,3466..3483,3484..3489))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3504..3525
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3642..3659)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3813..4401)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3893..3912)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4572..5363,5364..5432))
                     /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     5195..5214
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5433..5537)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    join(5567..5953,1..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 aaggacctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
      181 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
      241 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
      301 gtcagcgggt gttggcggcg cgccatcgag gagaacttct agtatatcta catacctaat
      361 attattgcct tattaaaaat ggaatcccaa caattacatc aaaatccaca ttctcttcaa
      421 aatcaattgt cctgtacttc cttgttcatg tgtgttcaaa aacgttatat ttataggata
      481 attatactct atttctcaac aagtaattgg ttgtttggcc gagcggtcta aggcgcctga
      541 ttcaagaaat atcttgaccg cagttaactg tgggaatact caggtatcgt aagatgcaag
      601 agttcgaatc tcttagcaac cattattttt ttcctcaaca taacgagaac acacaggggc
      661 gctatcgcac agaatcaaat tcgatgactg gaaatttttt gttaatttca gaggtcgcct
      721 gacgcatata cctttttcaa ctgaaaaatt gggagaaaaa ggaaaggtga gagcgccgga
      781 accggctttt catatagaat agagaagcgt tcatgactaa atgcttgcat cacaatactt
      841 gaagttgaca atattattta aggacctatt gttttttcca ataggtggtt agcaatcgtc
      901 ttactttcta acttttctta ccttttacat ttcagcaata tatatatata tatttcaagg
      961 atataccatt ctaatgtctg cccctaagaa gatcgtcgtt ttgccaggtg accacgttgg
     1021 tcaagaaatc acagccgaag ccattaaggt tcttaaagct atttctgatg ttcgttccaa
     1081 tgtcaagttc gatttcgaaa atcatttaat tggtggtgct gctatcgatg ctacaggtgt
     1141 tccacttcca gatgaggcgc tggaagcctc caagaaggct gatgccgttt tgttaggtgc
     1201 tgtgggtggt cctaaatggg gtaccggtag tgttagacct gaacaaggtt tactaaaaat
     1261 ccgtaaagaa cttcaattgt acgccaactt aagaccatgt aactttgcat ccgactctct
     1321 tttagactta tctccaatca agccacaatt tgctaaaggt actgacttcg ttgttgtcag
     1381 agaattagtg ggaggtattt actttggtaa gagaaaggaa gacgatggtg atggtgtcgc
     1441 ttgggatagt gaacaataca ccgttccaga agtgcaaaga atcacaagaa tggccgcttt
     1501 catggcccta caacatgagc caccattgcc tatttggtcc ttggataaag ctaatgtttt
     1561 ggcctcttca agattatgga gaaaaactgt ggaggaaacc atcaagaacg aattccctac
     1621 attgaaggtt caacatcaat tgattgattc tgccgccatg atcctagtta agaacccaac
     1681 ccacctaaat ggtattataa tcaccagcaa catgtttggt gatatcatct ccgatgaagc
     1741 ctccgttatc ccaggttcct tgggtttgtt gccatctgcg tccttggcct ctttgccaga
     1801 caagaacacc gcatttggtt tgtacgaacc atgccacggt tctgctccag atttgccaaa
     1861 gaataaggtc aaccctatcg ccactatctt gtctgctgca atgatgttga aattgtcatt
     1921 gaacttgcct gaagaaggta aggccattga agatgcagtt aaaaaggttt tggatgcagg
     1981 tatcagaact ggtgatttag gtggttccaa cagtaccacc gaagtcggtg atgctgtcgc
     2041 cgaagaagtt aagaaaatcc ttgcttaaaa agattctctt tttttatgat atttgtacat
     2101 aaactttata aatgaaattc ataatagaaa cgacacgaaa ttacaaaatg gaatatgttc
     2161 atagggtaga cgaaactata tacgcaatct acatacattt atcaagaagg agaaaaagga
     2221 ggatgtaaag gaatacaggt aagcaaattg atactaatgg ctcaacgtga taaggaaaaa
     2281 gaattgcact ttaacattaa tattgacaag gaggagggca ccacacaaaa agttaggtgt
     2341 aacagaaaat catgaaacta tgattcctaa tttatatatt ggaggatttt ctctaaaaaa
     2401 aaaaaaatac aacaaataaa aaacactcaa tgacctgacc atttgatgga gtttaagtca
     2461 ataccttctt gaaccatttc ccataatggt gaaagttccc tcaagaattt tactctgtca
     2521 gaaacggcct tacgacgtag tcgaggatcc tatgcggtgt gaaataccgc acagatgcgt
     2581 aaggagaaaa taccgcatca ggaaattgta agcgttaata ttttgttaaa attcgcgtta
     2641 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     2701 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     2761 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     2821 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     2881 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     2941 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc tggcaagtgt
     3001 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     3061 gtccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     3121 ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca
     3181 gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattgtaata cgactcacta
     3241 tagggcgaat tggagctcca ccgcggtggc ggccgctcta gaactagtgg gtcccccggg
     3301 ctgcaggaat tcgatatcaa gcttatcgat accgtcgacc tggagggggg gcccggtacc
     3361 agcttttgtt ccctttagtg agggttaatt tcgagcttgg cgtaatcatg gtcatagctg
     3421 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     3481 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     3541 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     3601 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     3661 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     3721 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     3781 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     3841 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     3901 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     3961 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     4021 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     4081 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     4141 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     4201 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta
     4261 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     4321 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     4381 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     4441 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     4501 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     4561 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     4621 cgttcatcca tagttgcctg actgcccgtc gtgtagataa ctacgatacg ggagggctta
     4681 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     4741 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     4801 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     4861 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     4921 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     4981 tgaaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     5041 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     5101 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     5161 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     5221 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     5281 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     5341 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     5401 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     5461 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     5521 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacggatc gcttgcctgt
     5581 aacttacacg cgcctcgtat cttttaatga tggaataatt tgggaattta ctctgtattt
     5641 atttattttt atgttttgta tttggatttt agaaagtaaa taaagaaggt agaagagtta
     5701 cggaatgaag aaaaaaaaat aaacaaaggt ttaaaaaatt tcaacaaaaa gcgtacttta
     5761 catatatatt tattagacaa gaaaagcaga ttaaatagat atacattcga ttaacgataa
     5821 gtaaaatgta aaatcacagg attttcgtgt gtggtcttct acacagacaa gatgaaacaa
     5881 ttcggcatta atacctgaga gcaggaagag caagataaaa ggtagtattt gttggcgatc
     5941 cccctagagt ctt
//
